SUP A OLEC LA R CAPSULES
Priority
This application claims priority to GB 1 2893.1 filed on 26 July 201 1 and GB 12021 27.5
filed on 08 February 2012, the contents of both of which are hereby incorporated by
reference in their entirety.
Fi&!d of the in ve tio
This invention relates to capsules, particularly microcapsules, based on a cucurbiturii
cross-linked network, and methods for the preparation of such capsules, and their use in
methods of delivering encapsulated components.
Background
The microencapsulation of a component by self-assembled hollow microspheres is one of
the important aspects of nanotechnology and materials science. Control over the shape
and composition of the supporting structure, parameters that influence the material
properties, is important for many applications, such as diagnostics, drug delivery,
electronic displays and catalysis (see Ke et al. Angew. Che 20 , 123, 3073; De Cock
et al. Angew. Chem. Int. Ed. 2010, 49, 6954; Yang et a l. Angew. Chem. 201 , 123, 497;
Comlskey et al. Nature 98, 394, 253; Peyratout et al. Angew. Chem. Int. Ed. 2004, 43,
3762). Preparation of conventional polymeric microcapsules proceeds via a iayer-bylayer
(L-b-L) scheme, where a solid support is coated by the sequential addition of a
series of oppositely charged poiyelectrolyte layers (see Caruso et a . Science 998, 282,
1 11; Donath et al. Angew. Chem. Int. Ed. 1998, 37, 2201 ) . This strategy provides a
uniform material but suffers from reduced encapsulation efficiencies due to the solid
template. An alternative method utilises colloidal emulsion-templating where liquid-liquid
interfaces drive the self-assembly of shell components (see C i et al. Adv. Fund Mater.
2010, 20, 1625). However, it is difficult to control monodispersity and material diversity of
the resulting microcapsules, thereby limiting its functionality in drug delivery and sensing
applications.
in contrast, microfiuidic droplets, a subset of colloidal emulsion, have shown great
promise for microcapsule fabrication (see Gunther et al. Lab Chip 2008, 6, 1487;
Huebner et al. Lab Chip 2008, 8, 1244; Theberge et al. Angew. Chem. Int. Ed. 2010, 49,
5846). These droplets of narrow size distribution (polydispersity index < 2%) can be
generated at extremely high frequency with economic use of reagents (see Xu et al.
AiChE Journal 2006, 52, 3005). Initial efforts to prepare capsules based on microdropletassisted
fabrication have focused on phase separation using double emulsion and liquid
crystal core temp!ating (see Utada et al. Science 2005, 308, 537; Priest et al. Lab Chip
2008, 8, 82). The formation of polymeric capsule walls has also been described in an
approach that involves microfiuidic device surface treatment and rapid polymerization
techniques (see Zhou et ai. Electrophoresis 2QQ , 31, 2; Abraham et a Advanced
Materials 2008, 20, 2177). The wall is formed as the solvent evaporates from formed
organic solvent droplets. Metal-organic framework capsules have also been recentlyreported
(see Ameloot et ai. Nat Chem. 20 , 3, 382). With the current ionic or covalent
cross-linking strategies, however, the main challenge in capsule fabrication lies in the
simultaneous production of uniform capsules with high cargo loading efficiencies and
facile incorporation of diverse functionality into the capsule shell.
The present inventors have now established a capsule based on a cucurbituri!-based
host-guest network. Designing microstructures using multivalency and cooperativity
through molecular recognition provides an unparalleled opportunity in the fabrication of
microcapsules with tailorabie interactions and functionalities. However, efforts in
preparing microcapsules using supramolecular host-guest approach, as described herein,
are scarce (see De Cock et al. Angew. Chem. int. Ed. 2010, 49, 8954).
Previous disclosures include a colloidal microcapsule comprising b-cyclodextrin and
modified gold nanoparticles (AuNPs) prepared via emulsion templating (Patra et aL,
Langmuir 2009, 25, 13852), and a microcapsule comprising polymers functionalized with
cyclodextrin and ferrocene prepared using a L-b-L synthesis (Wang et aL, Chemistry of
Materials 2008, 20, 4194).
Summary of the invention
The present invention generally provides capsules having a shell of material that is a
supramolecular cross-linked network. The network is formed from a host-guest
compiexation of cucurbiturii (the host) and one or more building blocks comprising
suitable guest functionality. The complex non-covalently crosslinks the building block
and/or non-covalently links the building block to another building block thereby forming
the network.
In a general aspect the present invention provides a capsule having a shell obtainable
from the compiexation of cucurbiturils with suitable guest molecules.
In a first aspect of the invention there is provided a capsule having a shell which is
obtainable from the compiexation of a composition comprising cucurbiturii and one or
more building blocks having suitable cucurbiturii guest functionality thereby to form a
supramolecular cross-linked network.
In one embodiment, the shell is obtainable from the compiexation of (a) a composition
comprising cucurbiturii and (1) or (2); or (b) a composition comprising a plurality of
covalently linked cucurbiturils and (1), (2) or (3).
In one embodiment, the shell is obtainable from the complexation of a composition
comprising cucurbituril and (1) or (2).
In one embodiment, the shell is obtainable from the complexation of a composition
comprising cucurbituril and ( 1 ) .
(1) comprises a first building block covaiently linked to a plurality of first cucurbituril guest
molecules and a second building block covaiently linked to a plurality of second
cucurbituril guest molecules, wherein a first guest molecule and a second guest molecule
together with cucurbituril are suitable for forming a ternary guest-host complex.
(2) comprises a first building block covaiently linked to a plurality of first cucurbituril guest
molecules and a plurality of second cucurbituril guest molecules, wherein a first and a
second guest molecule together with cucurbituril are suitable for forming a ternary guesthost
complex. Optionally the composition further comprises a second building block
covaiently linked to one or more third cucurbituril guest molecules, one or more fourth
cucurbituril guest molecules or both, wherein a third and a fourth molecule together with
cucurbituril are suitable for forming a ternary guest-host complex, and/or the first and
fourth molecules together with cucurbituril are suitable for forming a ternary guest-host
complex, and/or the second and third molecules together with cucurbituril are suitable for
forming a ternary guest-host complex;
(3) comprises a first building block covaiently linked to a plurality of first cucurbituril guest
molecules, wherein the first guest molecule together with the cucurbituril are suitable for
forming a binary guest-host complex. Optionally the composition further comprises a
second building block covaiently linked to one or more second cucurbituril guest
molecules, wherein the second guest molecule together with the cucurbituril are suitable
for forming a binary guest-host complex.
In one embodiment, the cucurbituril is selected from CB[8] and variants and derivates
thereof.
In one embodiment, the cucurbituril forms a ternary complex with suitable guest
molecules, for example with first and second guest molecules.
In one embodiment, the capsule is a microcapsule.
In one embodiment, the capsule encapsulates a component.
In a second aspect of the invention there is provided a method for the preparation of a
capsule having a shell, such as the capsule of the first aspect of the invention, the method
comprising the step of:
(i) contacting a flow of a first phase and a flow of a second phase in a channel,
thereby to generate in the channel a dispersion of discrete regions, preferably droplets, of
the second phase in the first phase, wherein the second phase comprises cucurbituril and
one or more building blocks having suitable cucurbituril guest functionality suitable to form
a supramoiecu!ar cross-linked network, thereby to form a capsule shell within the discrete
region, wherein the first and second phases are immiscible.
In one embodiment, the second phase comprises either (a) a cucurbituril and ( 1 ) or (2); or
(b) a plurality of covalentiy linked cucurbituri!s and ( ) , (2) or (3).
In one embodiment, one of the first and second phases is an aqueous phase and the
other phase is a water immiscible phase.
In one embodiment, the second phase is an aqueous phase. The first phase is a water
immiscible phase, for example an oil phase.
In one embodiment, the first phase is an aqueous phase. The second phase is a water
immiscible phase, for example an oil phase.
In one embodiment, the method further comprises the step of (ii) collecting the outflow
from the channel, thereby to obtain a droplet, which contains a capsule.
In one embodiment, the method comprises the step (ii) above and (iii) optionally drying
the capsule obtained in step (ii).
In one embodiment, the channel is a microfluidic channel.
In one embodiment, the flow of the second phase is brought into contact with the flow of
the first phase substantially perpendicular to the first phase in this embodiment, the
channel structure may be a T-junction geometry.
In one embodiment, the flow of the second phase further comprises a component for
encapsulation, and the step (i) provides a capsule having a shell encapsulating the
component.
In a third aspect of the invention there is provided a capsule obtained or obtainable by the
method of the second aspect of the invention.
In a fourth aspect of the invention there is provided a method of delivering a component to
a location, the method comprising the steps of:
(i) providing a capsule having a shell encapsulating a component;
(ii) delivering the capsule to a target location;
(iii) releasing the component from the shell.
In an alternative aspect of the invention there is provided a capsule having a shell which
is obtainable from the compiexation of a composition comprising a host and one or more
building blocks having suitable host guest functionality thereby to form a supramolecu!ar
cross-linked network.
in one embodiment the host is selected from cyciodextrin, calix[n]arene, crown ether and
cucurbituril, and the one or more building blocks having suitable host guest functionality
for the cyciodextrin, calix[n]arene, crown ether or cucurbituril host in one embodiment,
references in the first to fourth aspects above to cucurbituril and a cucurbituril guest may
be interpreted as a reference to an alternative host and a suitable guest for that host.
Summary of the Figures
Figure 1 (a) is a schematic representation of the microdroplet generation process using a
microfluidic flow focusing device, consisting of the oil continuous phase perpendicular to a
combination of aqueous solutions of CB[8] 1, MV +-AuNP 2, and Np-pol 3 as the
dispersed phase (b) Microscopic image and schematic representation of the flow
focusing region, with downstream mixing channel allowing thorough mixing of reagents
online (c) The high monodispersity of microfluidic droplets is demonstrated by its narrow
size distribution.
Figure 2 (a) Bright field images of the late stage of the capsule formation process as
water evaporates, rendering a collapsed microcapsule. Scale bar = 5 m h . (b) Light
microscope image of the exploded capsules showing the relics of the capsule shell.
Scale bar = 0 mhi . (c) SE image of a dried and at least partially collapsed capsule.
Scale bar = 2 m . (d) TE image of the microcapsule shell, showing 5 nm AuNPs
dispersed in a mesh of polymer. Scale bar = 0 nm. (e) Schematic representation of the
proposed microcapsule formation process from the initial droplet (with diameter d) to the
dehydrated stable capsule (with diameter d'). The cross-linking structure of 1 and 2 for the
capsule material is also proposed.
Figure 3 (a) Chemical structure and schematic representation of NP-RD-poi 4. (b) LSC
image of droplets containing aqueous solutions of Np-RD-pol, CB[8] and MV-AuNP, and
the fluorescence intensity profile. Scale bar = 40 m . (c) LSCM image of a droplet (46 mί
diameter) containing aqueous solutions of Np-RD-pol, CB[8], MV-AuNP and F!TC-dextran
and the corresponding fluorescence intensity profile. Scale bar = 7.5 mΐt i. (d) LSCM image
of a droplet (23 m diameter) containing aqueous solutions of Np-Rd-pol, CB[8],
MV-AuNP and FITC-dextran and the corresponding fluorescence intensity profile. Scale
bar = 10 m h .
Figure 4 is the bright field and fluorescence images of dried microcapsules containing
FITC-dextran before and after rehydration, showing (a) the expansion of the microcapsule
wail accompanied by the leakage of FITC-dextran ( 10 kDa), (b) the retaining of the
FITC-dextran (500 kDa), and (c) the partial permeability of the FITC-dextran (70 kDa) for
microcapsules containing twice concentrated CB[8] cross-linker. Scale bars = 20 mhi
Figure 5 (a) Schematic representation of the proposed effect of the reduction of V on
the ternary complex CB[8]:MV ÷-AuNP:Np-pol, and the resulting formation of 2:1
(MV *-AuNP) c:CB[8] complex (b) The fluorescence images of the process of the
disintegration of the microcapsule wall material in an aqueous solution of Na S 0 4 and in
H 0 over 12 h in N environment at 25 °C. Scale bars = = 5 m .
Figure 6 (a) Schematic representations of the microcapsules with and without
V +-AuNPs (5 nm and 20 nm). For negative control. V ÷-poi 5 was used instead of
AuNPs. (b) SERS spectra of empty microcapsules consisting of V +-pol, 5 nm
V +-AuNP, and 20 nm V +-AuNP, showing characteristic peaks for CB[8] and V +
(indicated with arrows) (c) SERS spectra of FITC-dextran encapsulated microcapsules
consisting of V2+-poi and 20 nm MV +-AuNP. showing characteristic peaks for FITC
(indicated with arrows) in addition to capsule shell materials. Ail spectra were obtained
using 633 nm excitation laser line (d) SERS mapping of the microcapsule, showing
localization of the SERS signal for CB[8] and V2+ .
Figure 7 is the excitation spectrum of Np-RD-pol and emission spectra excited at 514 nm
and 544 nm.
Figure 8 shows the variation in mean diameter of droplets as a function of the ratio of
Q i Q q , of various aqueous streams of Qaq = 80 m , 100 m i , 1 0 m 'h using T-junction
and a channel 40 mhi in width (solid line), and of various aqueous streams of Q =
40 m /h, 60 .L/h, 80 m using T-junction and a channel 20 m t in width (dashed line).
Figure 9 shows the variation in the mean diameter of droplets as a function of the ratio of
the oil and the aqueous stream, and as a function of the ratio of the individual aqueous
stream flow rates.
Figure 10 is the LSCM image of a droplet containing aqueous solutions of Np-RD-pol,
CB[8], V-AuNP and GFP-expressing £ . c / ceils and the corresponding fluorescence
intensity profile.
Detailed Description of the i ventio
The present inventors have established that capsules may be prepared having a shell that
is obtainable from the supramoleeular complexation of cucurbituril with building blocks
covalentiy linked to appropriate cucurbituril guest molecules.
The capsules are formed using fiuidic droplet generation techniques, amongst others.
The ability of cucurbituril and the building blocks to form a shell is surprising given the
previously reported behaviour of such materials.
Earlier work from one of the present inventors has found that cucurbituril may be used to
form a supramolecular cross-linked network via host-guest comp!exation (see Appe et al.
J. Am Che . Soc. 20 0, 132, 14251). The network is based on the supramolecular
assembly of a ternary complex of CB[8] together with a methyl viologen-functiona!ised
(MV) polymer and a napthol-functionaiised (Np) polymer. However, the networks
described here are in the form of supramolecular hydrogeis. Capsules are not described
or suggested.
The hydrogeis are prepared by sonication of the MV-functionalised polymer together with
CB[8], followed by addition of the Np-functionaiised polymer, with a subsequent short
mixing step.
The finding by the present inventors that cucurbituril can be mixed together with building
blocks connected to appropriate guest molecules thereby to yield a shell of material was
therefore unexpected. The capsule is obtainable through the use of fiuidic droplet
preparation techniques and bulk droplet generation techniques. The former is particularly
beneficial in that it generates droplets having a very low distribution of sizes, which results
in capsules having a very low distribution in sizes. Moreover, the methods of the
invention allow close control over the formation of the product capsule. Simple changes
in the fiuidic droplet preparation technique, such as changes in flow rates, may be used to
control the size of the capsule obtained, the size of the pores in the shell, and the
thickness of the shell, amongst others.
The capsules of the invention are shown to be robust, and are capable of withstanding
temperatures of at least I QO . The capsules also maintain their integrity at reduced
pressure.
The capsules of the invention are suitable for encapsulating a component. Using the
fiuidic droplet preparation techniques described herein, a capsule shell may be
constructed in the presence of the component to be encapsulated. Thus, in one
procedure the shell may be formed and the component encapsulated. Advantageously
therefore, the capsule may be constructed without the need for a later passive diffusion
step after the capsule construction. Furthermore, the method of encapsulation allows
high rates of incorporation of the material into the capsule, and material waste is therefore
minimised.
The invention is now described in more detail with reference to the each feature of the
invention.
Capsules
A capsule of the invention comprises a shell of material. The material is the
supramolecular complex that is formed from the complexation of cucurbituril with building
blocks covalently linked to appropriate cucurbituril guest molecule. The shell defines an
internal space, which may be referred to as a hollow space, which is suitable for holding a
component. Thus, in one embodiment, the capsules of the invention extend to those
capsules encapsulating a component within the shell. The shell may form a barrier
limiting or preventing the release of material encapsulated within.
The component may be reieasable from the capsule, through pores that are present in the
shell in some embodiments, the pores are sufficiently small to prevent the component
from being released. Thus, the network making up the shell may be at least partly
disassembled thereby permitting release of material from within the shell. Further pores
may be generated by the application of an external stimulus to the shell. In this case, the
pores may be generated through a disruption of the cucurbituril guest complex. Such
decompiexation therefore creates pores through which encapsulated components may be
released from within the shell. In some embodiments of the invention, the shell material
may subsequently be reformed by reassembly of the shell components.
In one embodiment, the capsule holds water within the shell. The water may be an
aqueous solution comprising one or more of the reagents that are for use in the
preparation of the supramolecular shell i.e. unreacted reagents. In one embodiment, the
aqueous solution comprises cucurbituril and/or ( 1 ) or (2) or (b) a plurality of cova!entiy
linked cucurbituriis and/or ( 1 ) , (2) or (3). Within the shell there may also be present a
network that is formed from the complexation of the reagents that have been used to
generate the shell.
Within the shell there may be provided an encapsulated material, which may be provided
in addition to water and the reagents that are for use in the supramolecular assembly of
the shell.
Where the capsule is said to encapsulate a component, it is understood that that this
encapsulated component may be present within the internal space defined by the shell.
In one embodiment, the encapsulant is also present, at least partially, within the pores of
the shell.
The presence of a component within the shell and/or within the pores of the shell may be
determined using suitable analytical techniques which are capable of distinguishing the
shell material and the encapsulant. For example, each of the shell material and the
component may have a detectable label or suitable functionality that is independently
detectable (orthogonal) to the label or functionality of the other. In one embodiment, each
of the shell and the component has an orthogonal fluorescent label. For example, one
has a rhodamine label and the other has a fluorescein label. Laser scanning confocal
microscopy techniques may be used to independently detect the fluorescence of each
label, thereby locating each of the shell and encapsulant. Where the component signals
are located at the same point as the signals from the shell, it is understood that the
component resides within a pore of the shell.
The general shape of the shell, and therefore the shape of the capsule, is not particularly
limited. In practice however, the shape of the capsule may be dictated by its method of
preparation. n the preparation methods described herein, a capsule shell may be
prepared using fluidic droplet formation techniques. Typically, the shell material is formed
at the boundary of a discrete (or discontinuous) phase in a continuous phase. For
example, one phase ma be an aqueous phase, and the other may be a water immiscible
phase. The discrete region may be a droplet, having a substantially spherical shape. The
shell formed is therefore also substantially spherical.
In certain embodiments, a capsule may be obtained when the shell has a substantially
spherical shape. This capsule may be subjected to a drying step, which reduces the
amount of solvent (for example, water) in and around the capsule. As a result of this step,
the capsule shrinks in size. At first the shell maintains a substantially spherical shape.
After further drying, the capsule sphere may partially of fully collapse in on itself. The
structural integrity of the capsule is maintained and the shell simply distorts to
accommodate changes in the internal volume. Thus, the capsules of the invention
include those capsules where the shell is an at least partially collapsed sphere.
Given the formation of the capsule shell at the boundary of the discrete region (for
example, a droplet), references to the dimensions of a droplet may also be taken as
references to the dimension of the capsule. The capsule shell may form prior to a drying
step.
The inventors have established that capsules that have been shrunk, for example by
desoivation, may subsequently be returned to their original substantially spherical shape,
by, for example, resolvating the capsule.
The shape of a capsule may be determined by simple observation of the formed capsule
using microscopy, such as bright field microscopy, scanning electron microscopy or
transmission electron microscopy. Where the shell material comprises a label, the
detection of the label through the shell will reveal the capsule shape. For example, where
the label is a fluorescent label, laser scanning confocal microscopy may be used to locate
the shell material and its shape.
The size of the capsule is not particularly limited. In one embodiment, the capsule is a
microcapsule and/or a nanocapsule.
In one embodiment, each capsule has an average size of at least 0.1 , 0.2, 0.5, 0.7, 1, 5,
10, 20, 30, 40, 50, 100 or 200 mhh in diameter.
In one embodiment, each capsule has an average size of at most 400, 200, 100, 75 or
50 mhi in diameter.
In one embodiment, the capsule size is in a range where the minimum and maximum
diamters are selected from the embodiments above. For example, the capsule size is in
range from 0 to 100 , m in diameter.
Average size refers to the numerical average of measured diameters for a sample of
capsules. Typically, at least 5 capsules in the sample are measured. A cross section
measurement is taken from the outmost edges of the shell.
The cross-section of a capsule may be determined using simple microscopic analysis of
the formed capsules. For example, the formed capsules may be placed on a microscope
slide and the capsules analysed. Alternatively, the capsule size may be measured during
the preparation process, for example as the capsules are formed in a channel of a fluidic
device (i.e. in line).
The measurement of the cross section may also be achieved using techniques related to
the detection of a detectable label or functionality present within the shell material. As
mentioned above in relation to detection and location of the encapsulated component, the
shell material may comprise a fluorescent label which may be detected by laser scanning
confocal microscopy techniques. The presence of multiple labels within and around the
capsule shell allows the cross-sectional shape to be determined, and the largest crosssection
measured.
In the preparation method described herein a capsule is prepared using a fluidic droplet
generation technique. The capsule shell is formed in a droplet, which is created in a
channel of a fluidic droplet generating device, at the boundary of the aqueous phase of
the droplet with the continuous phase. The size of the capsule is therefore substantially
the same as that of the droplet.
The present inventors have established that the capsules of the invention may be
prepared with a low size distribution. This is particularly advantageous, as a large
number of capsules ma be prepared, each with predictable physical and chemical
characteristics.
In one embodiment, the capsule diameter has a relative standard deviation (RSD) of at
most 0.5%, at most 1%, at most .5%, at most 2%, at most 4%, at most 5%, at most 7%,
or at most 10%.
The relative standard deviation is calculated from the standard deviation divided by the
numerical average and multiplied by 100. The size of the capsule refers to the largest
cross section of the capsule, in any section. The cross-section of a substantially spherical
capsule is the diameter.
The shell defines an interna! cavity which is suitable for encapsulating a component. The
size of the internal space will generally correspond to the size of the capsule itself. Thus,
the dimension, for example the diameter, of the internal space may be selected from any
one of the diameter values given above for the shell itself.
Where the size of the capsule is measured, the diameter refers to the distance from the
outermost edge to outmost edge of the shell material of two opposing points, as
mentioned above. Where the size of the internal space is measured, the diameter refers
to the distance from the innermost edge to innermost edge of the shell material of two
opposing points
The inventors have established techniques that allow the shell outer and inner edges to
be determined. For example, the presence of a detectable label within the shell material
allows the outermost and innermost edges of the shell to be determined if these edges
can be detected, the thickness of the shell may be determined.
Typically, the diameter as measured from outermost to outermost edge is not significantly
different to the diameter as measure from innermost to innermost edge. The difference is
the thickness of the shell at the two opposing points.
In one embodiment, the shell has a thickness of at least 0.02, at least 0.05, at least 0 . , at
least 0.5, at least .0, at least 2.0 or at least 5.0 m i.
As previously noted, the shell has pores. In one embodiment, the pores may be of a size
to permit the passage of material therethrough. For example, components encapsulated
within the capsule may pass through the pores of the shell to be released from the
capsule. Conversely, the pores may be of sufficient size to allow components to pass into
the shell internal space, and thereby become encapsulated. Such may be referred to as
a passive diffusion encapsulation step. Such a technique may be used to provide a
capsule having an encapsulant within. As described herein, the present inventors have
provided alternative methods for the encapsulation of material in the shell preparation
step. Such methods allow for a more efficient loading of the capsule with material, as the
material is entirely encapsulated within the shell.
In one embodiment, the pores may be of a size that is too small to permit passage of
material therethrough. For example, components encapsulated within the capsule may
be prevented from passing through the pores of the shell, and therefore cannot be
released from the capsule. Such material may be released from the capsule by, for
example, disrupting the cucurbituril complexes that hold the shell together. Disruption of
the shell in this way creates larger pores through which material may pass.
It is believed that the pore size may be increased upon solvation of a previously
desoivated capsule. As the capsule shrinks, the porosity of the capsule may decrease as
the shell material folds over, thereby at least partially blocking some of the pores.
The size of a pore may be gauged experimentally using a range of encapsulated
components each having a different cross-section, such as a different diameter. The
cross-section may be known or may be predicted based on an understanding of the likely
configuration of the component. The pore size may be determined based on which
components are released from the capsule and which are not.
The cross-section, typically diameter, of a component may be predicted based on the
calculated radius of gyration for each encapsulated component. Such calculations are
most suitable for determining the size of small globular particles, and may be used in
relation to polymeric systems, such as polypeptides, polynucleotides and
polysaccharides. Methods for the calculation of radius of gyration are described in
Andrieux et al. Analytical Chemistry 2002, 74, 5217, which is incorporated by reference
herein.
A capsule comprising an encapsulated component may be prepared using the methods
described herein. Once the capsule (with encapsulant) is prepared, the capsule and its
aqueous surroundings may be analysed for loss of material from within the shell out to the
external aqueous phase. The encapsulated compounds ma have an analytical label to
aid detection. Suitable labels include fluorescent labels which are detectable using
standard fluorescence microscopy techniques.
In one embodiment, dextran compounds of differing molecular weight may be used as
test compounds to determine the pore size of a formed capsule. The dextran may be
labelled, and preferably with a fluorescent label.
Dextran compounds of differing molecular weight are readily available from commercial
sources, including, for example, Sigma Aidrich. Dextrans having an average molecular
weight of from 1,000 to 500,000 are available. Dextran with a molecular weight of 70 kDa
has a radius of gyration of approx. 8 nm, whilst dextran with a molecular weight of
150 kDa has a radius of gyration of approx. 11 nm (see Granath Journal of Colloid
Science 1958, 13, 308). Dextran compounds having a fluorescent label, such as
fluorescein isothiocyanate, are also available from commercial sources, including, again,
Sigma Aidrich.
in one embodiment, the pore size is at most 20, at most 15, at most 0, at most 5, at most
1 or at most 0.5 m h
in one embodiment, the pore size is at most 500, at most 200, at most 100, at most 50, or
at most 20 nm.
In one embodiment, the pore size is at least 0.5, at least 1, or at least 5 nm.
in one embodiment, the pore size is in a range where the minimum and maximum pore
sizes are selected from the embodiments above. For example, the pore size is in range
1 to 20 nm.
As an alternative to dextran, protein standards may be used instead. As an alternative to
the labelled compounds described above, it also possible to detect the compound
released from the capsule using mass spectroscopy, or protein gel electrophoresis (for
protein standards).
Surface area, porosity and pore size may also be determined experimentally using BET
gas absorption techniques.
As expected, the shell pore size is influenced by the amount of cucurbituril present in the
complexable composition from which the capsule may be prepared increasing the
amount of cucurbituril present in the complexable composition is believed to increase the
amount of crosslinking with the network, thereby reducing the size of the pores in the
formed shell material.
The capsule shell may comprise one or more layers of material. The layers of the
material may be linked, for example by a ternary supramo!ecular complex of cucurbituril
with a first guest molecule present in one layer and a second guest molecule present in a
second layer. Additionally, or alternatively, the layers of the material may be linked by a
first building block having a plurality of guest molecules, where one guest molecuie forms
a ternary complex with a cucurbituril and another guest molecule present in a first layer,
and another guest molecule forms a ternary complex with a cucurbituril and another guest
molecule present in a second layer. In these embodiments the shell may be viewed as a
mesh extending in three dimensions. Although the shell may have a depth of material,
such as a thickness described herein, it is understood that the formation of the shell will
nevertheless provide an internal space in which a component may reside. Thus, the
present invention is not intended to encompass particles having no internal space.
Alternatively the capsule shell may comprise a plurality of concentric layers of network
material that are not interlinked. In any such embodiment, the reference to capsule size
refers to the cross section of the outermost shell.
As discussed above, the shell material may include detectable labels or detectable
functionalities
A detectable functionality is functionality of a capsule shell component having a
characteristic that is detectable over and above the characteristics that are present in
other components of the capsule, or even other functionalities of the same component.
The detectable functionality may refer to a particular chemical group that gives rise to a
unique signal in, for example, R, UV-ViS, NMR or Raman analysis. The functionality
may be a radioactive element.
Typically a part of the shell material or the encapsulant is provided with a detectable label,
as the introduction of a chosen label allows the use of techniques that are most
appropriate for the property that is to be measured. Described herein are building blocks
having fluorescent detectable labels. Also described herein are building blocks that are
capable of providing a surface enhanced resonance effect.
The shell may have additional functionality on its inner and/or outer surfaces. Described
herein are building blocks having functionality to improve solubility, to aid detection,
reactive functionality for later elaboration of the shell, and catalysts, amongst others.
The capsule shell of the invention is stable and may be stored without loss of the shell
structure. The integrity of the shell therefore allows the capsule to be used as a storage
vessel for an encapsulant. The capsules of the invention are thermally stable and the
shell is known to maintain its integrity at least up to 100°C. The capsules of the invention
are also stable at reduced pressures (i.e. below ambient pressure). The shell is known to
maintain its integrity down to at least 20 Pa.
The capsules of the invention have a long shelf life. The present inventors have
confirmed that structural integrity is maintained for at least 0 months.
The structural integrity of the shell is in part due to the strength of the cucurbiturii guesthost
complex, which is described in more detail below.
Additional or Alternative Capsule Features
The capsule shell has pores. The porosity is adjustable by appropriate changes in the
stoichiometry of the reagents used to form the capsule. Increasing the crossiinking
between building blocks will decrease the size of the pores in the capsule. Alternatively,
the building blocks may be selected so as to provide a shell material that has increased or
decrease porosity. Where a encapsulant or relatively small size is to be encapsulated,
the capsule is prepared with pores of relatively small diameter, thereby to limit or prevent
loss of the encapsulant out of the shell. Where a relatively large encapsulant is to be
encapsulated, the pore size may be larger.
As noted above, the shell may have additional functionality on its inner and/or outer
surfaces in some embodiments, the functionality is provided for later chemical
functionalisation of the capsule shell, for example as a reaction site for linking to a
compound having a particularly desirable reactivity.
In one embodiment, the shell has a chemical functional group available for reaction on the
outer and/or the inner surface of the capsule. The chemical functional group is selected
from the group consisting of hydroxy!, amine (preferably primary and secondary amine),
carboxy, thiol, ester, thioester, carbonate, urethane, and thiourea.
In one embodiment the shell is linked to a functional compound.
In one embodiment, the functional compound is an analytical label to aid detection and
quantification of the capsule. Such is described in the section above.
The functional compound may be catalytic (including enzymatic), antifungal, herbicida!, or
antigenic.
The functional compound may have surface adhesion properties. Such functionality may
be used to attach the capsule to a surface, either covalentiy or non-covalently.
The functional compounds may be capable of binding to (or sequestering) a compound or
ion. Such functionality may be of assistance in purification, such as filtering, and for the
capture of toxic and non-toxic elements and compounds.
In one embodiment, the functional compound is a biomoiecule.
In one embodiment, the functional compound is a polypeptide, a polysaccharide, a
polynucleotide, or a lipid.
Examples of polypeptides include enzymes, antibodies, hormones and receptors.
The functionality may be introduced into the shell by appropriate choice of building block
material. Thus, where the building block is a polymer, suitable functionality may be
incorporated into the monomers of the polymer, which monomer may be present within
the polymer backbone, or on a side chain. Where the building block is a particle, the
surface of that particle may be suitably functionalised.
Where a functional molecule is present on a surface of the shell, this molecule may be
added after the capsule is formed. The functional molecules may be linked to the shell
using a chemical functionai group that has been introduced for this purpose.
In principle, the cucurbituril may have functionality that is available for reaction. However,
this is less preferred.
Where necessary, appropriate protecting groups may be used to protect the functionality
during the capsule preparation procedure. The protecting groups may be removed later,
as and when required.
Complex
The capsule shell comprises a network that is held together by a supramolecular
handcuff. The complex that forms this supramolecular handcuff is based on a cucurbituril
hosting one guest (binary complex) or two guests (ternary complex). The cucurbituril
forms a non-covalent bond to each guest. The present inventors have established that
complexes of cucurbituril are readily formed and provide robust non-covalent linkages
between building blocks. The formation of the complex is tolerant of many functionalities
within the building blocks. One of the present inventors has demonstrated that polymer
networks may be prepared using a cucurbituril handcuff. However, until now, the
formation of precise polymer structures, such as capsules, using cucurbituril has been
described.
In one embodiment, the shell is a network having a plurality of complexes wherein each
complex comprises cucurbituril hosting a first guest molecule and a second guest
molecule. The first and second guest molecules are covalently linked to a first building
block, or to a first building block and a second building block.
Where the complex comprises two guests within the cucurbituril cavity, the association
constant, Ka, for that complex is at least 103 M2 , at least 04 2 , at least 05 2, at least
106 M2, at least 07 2 , at least 10s M2 , at least 109 2 , at least 0 0 2, at
least 101 1 2, or at least 1012 2.
Where a cucurbituril hosts two guest molecules, the guest molecules may be the same or
they may be different. A cucurbituril that is capable of hosting two guest molecules may
also be capable of forming a stable binary complex with a single guest. The formation of
a ternary guest-host complex is believed to proceed via an intermediate binary complex.
Within the shell, there may be present a binary complex formed between a guest
molecule and a cucurbituril. The binary complex may be regarded as a partially formed
ternary complex that has not yet formed a non-covaient bond to another guest molecule.
In one embodiment, the shell is a network having a plurality of complexes, wherein each
complex comprises cucurbituril hosting one guest molecule, and each cucurbituril is
covalently linked to at least one other cucurbituril. The guest molecules are covalently
linked to a first building block, or to a first building block and a second building block.
Where the complex comprises one guest within the cucurbituril cavity, the association
constant, K , for that complex is at least 103 of at least 104 , of at least 105 ~ , of
at least 06 M of at least 07 , of at least 108 M of at least 09 M , of at
least 1010 M , of at least 0 M , or of at least 0 12 M .
In one embodiment, the guest is a compound capable of forming a complex which has an
association constant in the range 104 to 0 M .
In one embodiment the formation of the complex is reversible. The decomplexation of the
complex to separate the guest or guests may occur in response to an external stimulus,
including, for example, a competitor guest compound. Such decomplexation may be
induced in order to provide additional or larger pores in the capsule through which an
encapsulated material may pass.
As noted above in relation to the capsule shell, the complex of cucurbituril with one or two
guests is the non-covaient link that links and/or interlinks the building blocks to from a
supramoiecu!ar network of material. The complex is thermally stable and does not
separate at reduced pressure, as explained for the shell.
Network
The formation of a supramoiecu!ar complex serves to link and/or interlink building blocks,
thereby forming a network of material. This is the capsule shell.
Two types of network are provided. The first type is based on the formation of a plurality
of ternary complexes, each complex comprising a cucurbituril host with a first guest
molecule and a second guest molecule. The second type is based on the formation of a
plurality of binary complexes, each complex comprising a cucurbituril host with a first
guest molecule. In this second type, each cucurbituril is covalently linked to a least one
other cucurbituril. These types of network may be combined with a shell.
Where a building block is provided with a plurality of guest molecules, all of the guest
molecules need not participate in a complex with cucurbituril. Where the network is
based on linking between ternary structures, a guest molecule of a building block may be
in a binary complex with a cucurbituril. The binary complex may be regarded as a
partially formed ternary complex that has not yet combined with a further guest molecule
to generate the ternary form.
Throughout the description references are made to a building block, a first building block
and a second building block. It is understood that a reference to such is a reference to a
collection of the individual molecules, particles, polymers etc. that are the building blocks.
Where a reference is intended to an individual building block molecule, particle etc. the
term single is used in reference to the building blocks e.g. a single first building block.
The networks described below are the basic networks that are obtainable from the
compositions described it is understood that the present inventions extends to more
complex networks that are obtainable from compositions comprising further building
blocks.
Network of Ternary Complexes Based on Cucurbituril
This network is obtainable from the assembly of a first guest molecule and a second
guest molecule together with a cucurbituril host. The guest molecules may be provided
on one or two (or more) building blocks as described below.
In one embodiment, a network is obtainable or obtained from the complexation of a
composition comprising a cucurbituril, a first building block covalently linked to a plurality
of first cucurbituril guest molecules and a second building block covalently linked to a
plurality of second cucurbituril guest molecules, wherein a first guest molecule and a
second guest molecule together with cucurbituril are suitable for forming a ternary guesthost
complex.
The ternary complex serves to non-covalentiy link the first and second building blocks. A
single first building block may form a plurality of non-covalent links to a plurality of second
building blocks. Similarly, a single second building block may form a plurality of noncovalent
links to a plurality of first building blocks. In this way, a network of material is
established.
It is noted that in some embodiments, the first and second guest moiecuies may be
identical. Therefore the first and second building blocks may differ in their compositions.
In some embodiments, the first and second building blocks may be identical. In this case,
the first and second guest molecules are different.
Shown below is a schematic structure of a basic network formed between cucurbituril, a
single first building block and two single second building blocks in the schematics
included in this text, the guest molecules are depicted as rectangles which are covalently
linked (vertical line) to a building block (horizontal line). The vertical line may depict a
direct covalent bond or a linker to the building block. The building block may be a
polymeric molecule, a particle or the like, as described herein.
In the schematic below, some of the first guest moiecuies (unshaded rectangles) of the
first building block are in complex with cucurbituril hosts (barrels) and second guest
molecules (shaded rectangles) of the second building blocks.
Second Second
It is apparent that not all guest moiecuies present participate in a complex in the final
network. Each of the first and second building blocks may form complexes with other
second and first building blocks respectively. The guest molecules are shaded for ease of
understanding. However, as explained herein, the guest moiecuies of the first and
second building blocks may be the same.
In an alternative embodiment, a network is obtainable or obtained from the complexation
of a composition comprising a cucurbituril and a first building block covalently linked to a
plurality of first cucurbituril guest molecules and a plurality of second cucurbituril guest
molecules, wherein a first and a second guest molecule together with cucurbituril are
suitable for forming a ternary guest-host complex.
The ternary complex serves to non-covalently link and/or interlink the first building block.
A single first building block may form a plurality of non-covaient links to a plurality of other
first building blocks. Additionally, or alternatively, a single first building block may form a
plurality of non-cova!ent interlinks with itself, thereby to crosslink the single first building
block.
As before, the first and second guest molecules may be identical.
Shown below is a schematic structure of a basic network formed between cucurbituril and
two single first building blocks each having a plurality of first and second guest molecules.
Some of the first guest molecules (unshaded rectangles) of the first building block are in
complex with cucurbituril hosts (barrels) and second guest molecules (shaded rectangles)
of another first building block it can be seen from the network illustrated that a first
building block may form intramolecular complexes, thereby cross!inking a single first
building block.
First
It is apparent that not all guest molecules present need participate in a complex in the
final network. Each of the first building blocks may form complexes with other first
building blocks, or with other parts of the same building block. As explained herein, the
first and second guest molecules may be the same.
Optionally, the composition further comprises a second building block covalently linked to
one or more third cucurbituril guest molecules, one or more fourth cucurbituril guest
molecules or both, wherein a third and a fourth molecule together with cucurbituril are
suitable for forming a ternary guest-host complex, or the first and fourth guest molecules
together with cucurbituril are suitable for forming a ternary guest-host complex, or the
second and third guest molecules together with cucurbituril are suitable for forming a
ternary guest-host complex.
Where the second building block is provided with a plurality of third and fourth guest
molecules, the ternary complex serves to non-covalently link and/or interlink the second
building block. A single second building block may form a plurality of non-covalent links to
a plurality of other second building blocks. Additionally, or alternatively, a single second
building block may form one or more non-covalent interlinks with itself, thereby to
crosslink the single second building block.
The third and fourth guest molecules may be suitable for forming complexes with the first
and second guest molecules of the first building block. In one embodiment, the first and
third guest molecuies are the same. In one embodiment the second and fourth guest
molecules are the same. Here, the ternary complex serves to non-covalently link the first
and second building blocks, for example through a complex of the first and fourth guest
molecuies and/or through a complex of the second and third guest molecuies.
Thus, a single first building block may form a plurality of non-covalent links to a plurality of
second building blocks. Similarly, a single second building block may form a plurality of
non-covalent links to a plurality of first building blocks. In this way, a network of material
is established. The building blocks may also form intermoiecu!ar non-covalent bonds as
described previously.
Where a second building block is covaiently linked to one or more third guest molecules
or one or more fourth guest molecule, the first and fourth molecules together with
cucurbituril are suitable for forming a ternary guest-host complex, and the second and
third molecules together with cucurbituril are suitable for forming a ternary guest-host
complex. Thus, the ternary complex serves to non-covalently link the second building
block to the first building block.
Shown below is a schematic structure of a basic network formed between cucurbituril,
three single first building blocks each having a plurality of first and second guest
molecules, and two second building blocks each having a plurality of third and fourth
guest molecules. Some of the first guest molecules (unshaded rectangles) of the first
building block are in complex with cucurbituril hosts (barrels) and second guest molecules
(shaded rectangles) of another first building block. Some of the third guest molecules
(partially shaded rectangles) of the second building block are in complex with cucurbituril
hosts (barrels) and fourth guest molecules (dashed rectangles) of another second building
block. A the first guest molecule of the first building block is in complex with a cucurbituril
host and a fourth guest molecule (dashed rectangles) of a second building block. A
second guest molecule of the first building block is in complex with a cucurbituril host and
a third guest molecule of a second building block.
First Second
First
The first and third guest molecules may be the same. The second and fourth guest
molecules may be the same.
A second building block may be covalently linked to one guest molecule (which may be a
third or a fourth guest molecule). n this embodiment, the second building block is not
capable of forming a plurality of links to other building blocks. As such, the building block
would not contribute to the formation of a cross links within the network. However, the
second building block may be provided in order to introduce into the network a particular
physical or chemical characteristic that is possessed b the second building block. For
example, the second building block may comprise a detectable label or a functional
group, such as a solubiiising group. The incorporation of the second building block into
the network therefore allows the modification of the physical or chemical characteristics of
the overall network.
Shown below is a schematic structure of a basic network formed between cucurbituril, two
single first building blocks each having a plurality of first and second guest molecules, and
also including a single second building block, which is covalently linked to one fourth
guest molecule, and a detectable label. Some of the first guest molecules (unshaded
rectangles) of the first building block are in complex with cucurbituril hosts (barrels) and
second guest molecules (shaded rectangles) of another first building block. A first guest
molecule of the first building block is in complex with a cucurbituril host and a fourth guest
molecule. The detectable label (partially shaded circle) may be provided in order to allow
identification of the resulting network.
First Second
Network of Binary Complexes Based on a Plurality of Covalently Linked Cucurbiturils
This network is obtainable from the assembly of a first guest molecule together with a
cucurbituril host, which host is covalently linked to one or more other cucurbiturils. The
guest molecules may be provided on one, or two (or more) building blocks as described
herein.
The covalently linked cucurbiturils serve to link building block molecules through the
plurality of complexes that are formed within each of the covalently linked cucurbiturils.
Shown below is a schematic structure of a basic network formed between a plurality of
covalently linked cucurbiturils and two single first building blocks each having a plurality of
first guest molecules. Some of the first guest molecules (unshaded rectangles) of each of
the single first building block are in a binary complex with cucurbituril hosts (barrel). The
cucurbiturils are linked, thereby to form a link between each of the first building blocks.
Fi st
It is apparent that not all guest molecules present need participate in a complex in the
final network. Each of the single first building blocks may form complexes with other first
building blocks respectively, or may form an intramolecular crosslink with another portion
of the same building block. As explained herein, the guest molecules of the first and
second building blocks may be the same. In the schematic above, one of the fist building
blocks may be replaced with a second building block which is covalently linked to a
second guest molecule. The second guest molecule is one that is capable of forming a
binary compiex with the cucurbituri!. The second guest molecule may be the same as the
first guest molecule.
In the schematic two cucurbiturils are shown linked together. The present invention
encompasses the use of systems where more than two cucurbiturils are linked together.
For example multiple cucurbiturils may be pendant to a polymeric molecule.
Network of Ternary Complexes Based on a Plurality of Covalentiy Linked Cucurbiturils
It will be apparent from the description of the networks above, that each of the cucurbituri!
hosts n the plurality of covalentiy linked cucurbiturils may be suitable for forming ternary
complexes. Thus, the plurality of covalentiy linked cucurbiturils may be used in place of
the cucurbituri! described for use in the network of ternary complexes based on
cucurbituri!.
Shown below is a structural schematic of a basic network formed between a plurality of
covalentiy linked cucurbiturils, two single first building blocks each having a plurality of
first guest molecules, and two single second building blocks each having a plurality of
second guest molecules. Some of the first guest molecules (unshaded rectangles) of the
first building block are in tertiary complex with a cucurbituri! host (barrel) and the second
guest molecules (shaded rectangles) of the second building block The cucurbiturils are
linked, thereby to form a link between each of the first and second building blocks.
As before, the first and second guest molecules may be the same. Each of the first and
second building blocks may form complexes with other second and first building blocks
respectively. Other permutations are possible, for example, where the plurality of
covalentiy linked cucurbiturils has greater than two cucurbiturils.
Other Networks
Described above are the basic networks of the invention that are obtained or obtainable
from the compositions described it will be clear to one of skill in the art that the
compositions described may include further building blocks, for example third and fourth
building blocks, each linked to one or more cucurbituri! guest molecules. The present
invention also covers capsules where the shell comprises a mixture of any one of the
networks described above. Such are obtainable from compositions comprising an
appropriate selection of cucurbituril, covaiently linked cucurbiturils, first building block and
second building block as appropriate.
The invention also relates to a capsule having a shell that is a network comprising
different cucurbiturils. Different cucurbiturils may be chosen in order to obtain a network
that is based on ternary and binary complexes. Different cucurbiturils may be chosen in
order to generate networks that result from the selective complexation of each cucurbituril
for different guest molecules, which may be present on the same or different building
blocks.
Cucurbituril
The present invention provides use of cucurbituril as a supramolecu!ar handcuff to link
and/or crosslink building blocks. The cucurbituril may be used to form ternary complexes
with first and second guest molecules present on one or more building blocks. The
formation of such complexes links individual building blocks thereby to form a network of
material. This network is the shell of the capsule.
Additionally, or alternatively, a plurality of covaiently linked cucurbiturils is provided and
each cucurbituril may be used to form binary complexes with a guest molecule present on
one or more building blocks. The formation of a binary complex with each of the
covaiently linked cucurbiturils thereby forms a network of material. This network is the
shell of the capsule.
In one embodiment, the cucurbituril is capable of forming a ternary complex. For
example, CB[8], is capable of forming a ternary complex.
In one embodiment, the cucurbituril is capable of forming a binary complex. For example,
CB[7], is capable of forming a binary complex.
In one embodiment, the cucurbituril is capable of forming ternary and binary complexes.
For example, CB[8], is capable of forming a ternary or a binary complex, depending upon
the nature of the guest.
In one embodiment, the cucurbituril is a CB[5], CB[6], CB[7], CB[8], CB[9], CB[10], CB[1 1]
or CB[ 2] compound.
In one embodiment, the cucurbituril is a CB[8], CB[7], or CB[8] compound.
In one embodiment, the cucurbituril is a CB[8] compound.
In one embodiment, references to a cucurbituril compound are references to variants and
derivatives thereof.
Cucurbituril compounds differ in their water solubility. The methods of capsule
preparation may be adapted to take into account this solubility, as described later.
Therefore the choice of cucurbituril compound is not limited by its aqueous solubility.
In one embodimeni, the cucurbitunl compound has a solubility of at least 0.01 mg/mL, at
least 0.02 mg/mL, at least 0.05 mg/mL, or at least 0.10 mg/mL.
In one embodiment, the solubility refers to aqueous solubility (i.e. an aqueous phase).
In one embodiment, the solubility refers to solubility in a water immiscible phase, such as
an oil phase or an organic phase.
Cucurbit[8]uri! (CB[8]; CAS 259886-51-6) is a barrel shaped container molecule which has
eight repeat glycoluril units and an internal cavity size of 479A 3 (see structure below).
CB[8] is readily synthesised using standard techniques and is available commercially (e.g.
Siama-Aldrich, MO USA).
In other aspects of the invention, CB[8] variants are provided and find use in the methods
described herein.
A variant of CB[8] may include a structure having one or more repeat units that are
structurally analogous to glycoluril. The repeat unit may include an ethylurea unit. Where
all the units are ethylurea units, the variant is a hemicucurbituril. The variant may be a
hemicucurbit[12]uril (shown below, see also Lagona et al. Angew. Chem. Int. Ed. 2005,
44, 4844).
In other aspects of the invention, cucurbiturii derivatives are provided and find use in the
methods described herein. A derivative of a cucurbiturii is a structure having one, two,
three, four or more substituted glycoluril units. A substituted cucurbiturii compound may
be represented by the structure below:
wherein:
n is an integer of at least 5;
and for each glycoluril unit
each X s O, S or NR3, and
-R and -R2 are each independently selected from -H and the following
optionally substituted groups: -R3, -OH, -OR3, -COOH, -COOR3, -NH , -NHR3 and -N(R )2
where -R3 is independently selected from d -2oalkyl, C6-2ocarboaryl, and G 0heteroary!, or
where -R and/or -R2 is -N(R )2, both -R3 together form a C 7 heterocyclic ring; or together
-R and -R2 are C^alkylene forming a C6-Scarbocyclic ring together with the uracil frame.
In one embodiment, one of the giycolurii units is a substituted giycoluri! unit. Thus, -R
and -R2 are each independently -H for n-1 of the giycolurii units
In one embodiment, n is 5, 6, 7, 8, 9, 0, 1 or .
In one embodiment, n is 5, 6, 7, 8, 10 or 1 .
In one embodiment, n is 8.
In one embodiment, each X is O.
In one embodiment, each X is S.
In one embodiment, R and R2 are each independently H.
In one embodiment, for each unit one of R and R2 is H and the other is independently
selected from -H and the following optionally substituted groups: R3, -OH, -OR3, -COOH,
-COOR3, -NH , -NHR3 and -N(R ) . n one embodiment, for one unit one of R and R2 is H
and the other is independently selected from -H and the following optionally substituted
groups: -R3, -OH, -OR3, -COOH, -COOR3, -NH2 > -NHR3 and -N(R } . in this embodiment,
the remaining giycolurii units are such that R and R2 are each independently H.
Preferably -R3 is C 20alkyl, most preferably d 6aikyi. The C =:
a y group ma be linear
and/or saturated. Each group -R3 may be independently unsubstituted or substituted.
Preferred substituents are selected from: -R4, -OH, -OR4, -SH, -SR4, -COOH, -COOR" ,
-NH , -NHR" and -N R } , wherein -R4 is selected from a , C6- ocarboaryi, and
C5-2oheteroaryi. The substituents may be independently selected from -COOH and
-COOR4.
In some embodiments, -R4 is not the same as -R3. in some embodiments, -R is
preferably unsubstituted.
Where -R and/or -R2 is -OR3, -NHR3 or -N(R ) , then -R3 is preferably d -6alkyl. in some
embodiments, -R3 is substituted with a substituent -OR4, -NHR4 or -N(R ) . Each -R4 is
d-ealkyl and is itself preferably substituted
In some embodiments of the invention there is provided the use of a plurality of covalently
linked cucurbiturils. Such covalently linked cucurbiturils are suitable for forming networks
based on the complexation of the cucurbituril with guest molecules of a building block.
The complexes formed may be ternary or binary complexes.
A cucurbituril may be covalently linked to another cucurbituril via a linker group that is a
substituent at position R or R2 at one of the glycoluril units in the cucurbituril as
represented in the structure shown above. There are no particular limitations on the
covalent link between the cucurbiturils. The linker may be in the form of a simple alkylene
group, a polyoxyalkylene group or a polymer, such as a polymeric molecule described
herein for use in the building block. Where the linker is a polymeric molecule, the
cucurbiturils may be pendant to that polymer.
Building Block
Cucurbituril is used as a supramo!ecular handcuff to join together one or more building
blocks. The formation of a complex of the cucurbituril with suitable guest components
that are linked to the building blocks forms a network of material. This material is the
capsule shell. The complex non-covaiently crosslinks the building block or non-covalently
links the building block to another building block.
It is understood from the above that a building bock is an entity that serves to provide
structure to the formed network. The building block also serves as the link between a
plurality of guest molecules, and it may therefore also be referred to as a linker. In some
embodiments, a building block is provided for the purpose of introducing a desirable
physical or chemical characteristic into the formed network. As mentioned above in
relation to the network, a building block may include a functionality to aid detection and
characterisation of the shell. Such building blocks need not necessarily participate in a
crosslink.
A building block, such as a first building block, may be covalently linked to a plurality of
cucurbituril guest molecules. A building block will therefore non-covalently link to a
plurality of cucurbiturils, which cucurbiturils will non-covaiently link to other building
blocks, thereby to generate a network of material.
A building block, such as a first building block or a second building block, may be
covalently linked to a plurality of cucurbituril guest molecules. In one embodiment, a
building block is covalently linked to at least 3, at least 4, at least 5, at least 0, at least
20, at least 50, at least 100, at least 500, at least 1,000, at least 2,000, at least 5,000 or at
least 10,000 cucurbituril guest molecules.
In certain embodiments, building blocks covalently linked to one or more cucurbituril guest
molecules may be used. However, such building blocks are used only in combination
with other building blocks that are covalently linked to at least two cucurbituril guest
molecules.
In one embodiment, there is provided a first building block covalently linked to a plurality
of first guest molecules and a second building block covalently linked to a plurality of
second guest molecules. Each of the first and second building blocks may be covalently
linked to at least the number of guest molecules described above.
In one embodiment, there is provided a first building block covalently linked to a plurality
of first guest molecules and covalently linked to a plurality of second guest molecules.
The first building block may be covalently linked to at least the number of guest molecules
described above, which numbers may refer independently to the number of first guest
molecules and the number of second guest molecules.
In one embodiment, there is provided a second building block covalently linked to one or
more third guest molecules and/or covalently linked to a one or more fourth guest
molecules. In one embodiment, the second building block is covalently linked to at least
the number of guest molecules described above, which numbers may refer independently
to the number of third guest molecules and the number of fourth guest molecules. Such a
second building block may be used together with the first building block described in the
paragraph above.
Throughout the description, references are made to first and second building blocks. In
some embodiments, the first and second building blocks may be distinguished from each
other owing to differences, at least, in the structure of the building blocks themselves. In
some embodiments, the structures of the first and second building blocks are the same.
In this case, the building blocks may be distinguished from each other owing to
differences, at least, in the guest molecules that are covalently linked to each of the first
and the second guest molecules. Thus the terms first and second are intended to convey
a difference between the first building block together with its guest molecules and the
second building block together with its guest molecules.
The building blocks are not particularly limited, and the building block includes
compounds and particles, and may encompass assemblies of either of these. The guest
molecules are covalently linked to some portion of the building block.
At its simplest a building block is a linker for the connection of guest molecules.
In one embodiment the building block is a polymeric molecule or a particle.
Advantageously, a building block may be provided with certain functionality to aid the
formation of the capsule shell, or to improve its physical or chemical properties.
In one embodiment, the building block is provided with functionality to alter, or preferably
improve, water solubility. The functionality may take the form of a soiubiiising group, such
as a group comprising polyethylene glycol functionality. Other examples include groups
comprising amino, hydroxy, thiol, and carboxy functionality.
In one embodiment, the building block is provided with functionality to aid detection or
analysis of the building block, and to aid detection or analysis of the formed shell.
Advantageously, such functionality may also aid the detection of material encapsulated
within the shell. The functionality may take the form of a detectable label, such as a
fluorescent label.
In one embodiment, the building block is provided with reactive functionality for use in the
later elaboration of the shell material. The reactive functionality may be protected for the
shell forming reactions, then later deprotected to reveal the functionality. The functionality
may be a group comprising amino, hydroxy, thiol, and carboxy functionality.
Where the building block is provided with reactive functionality is provided, this
functionality may be suitable for linking the building block (and therefore the formed
capsule) to a surface.
In one embodiment, the building block is provided with a catalyst for later use in the
catalysis of a reaction at or near the shell surface. The catalyst may be provided at the
inner or outer edges of the shell thereby to catalyse internal and/or external reactions.
In one embodiment, the building block is chosen for its ability to influence the
opticoeiectronic properties of the encapsulant. Additionally or alternatively, the building
block may be chosen for its ability to be influenced by the encapsulant. The building
block may be suitable for transferring signals from the encapsulant to outside
environment.
In one embodiment a building block is capable of providing a surface enhanced
resonance effect.
Where functionality is provided it may be located at the outer side of, the inner side of
and/or within the shell. Thus, the functionality may be provided in connection with the
improvements related to the environment outwith the shell, within the internal space (the
space for holding an encapsulant) of the shell and/or within the shell (within the network of
shell material).
For the purposes of the methods described herein, the building block, together with the
guest molecules to which it is covaiently linked, should be soluble, for example in the
second phase.
In one embodiment, the building block has a solubility of at least 0.01 mg/mL, at least
0.02 mg/mL, at least 0.05 mg/mL, or at least 0.1 0 mg/mL.
In one embodiment, the solubility refers to aqueous solubility (i.e. an aqueous phase).
In one embodiment, the solubility refers to solubility in a water immiscible phase, such as
an oil phase or an organic phase.
A building block is linked to a cucurbituri! guest molecule or guest molecules by covalent
bonds. The covalent bond may be a carbon-carbon bond, a carbon-nitrogen bond, a
carbon-oxygen bond. The bond may be part of a linking group such as an ester or an
amide, and/or part of a group comprising an a!ky!ene or alkoxylene functionality.
Each guest molecule may be linked to the building block using routine chemical linkage
techniques. For example, guest molecules may be linked to the building block by:
aikyiation of a building block bearing an appropriate leaving group; esterification
reactions; amidation reactions; ether forming reactions; olefin cross metathesis; or small
guest molecule initiated reactions in which a polymer chain is grown off an initiating guest
molecule.
In one embodiment, the average molecular weight of a building block, optionally together
with any guest molecules, is at least 1,000, at least 5,000, at least 10,000, or at least
20,000.
In one embodiment, the average molecular weight of a building block, optionally together
with any guest molecules, is at most 30,000, at most 50,000, at most 100,000, at most
200,000, at most 500,000, at most ,000,000, or at most 2,000,000.
The average molecular weight may refer to the number average molecular weight or
weight average molecular weight.
In one embodiment, the average molecular weight of a building block is in a range where
the minimum and maximum amounts are selected from the embodiments above. For
example, the average molecular weight is in the range 1,000 to 100,000.
In one embodiment, a building block is capable of providing a surface enhanced
resonance effect. Typically, such capability is provided by a particle, and most particularly
a metal-containing particle. Suitable particles are such as those described herein. Most
suitable are those particles that are capable of providing a surface enhanced effect for
surface enhanced Raman spectroscopy.
Described below are building blocks that are based on polymeric molecules and particles,
including nanoparticies.
In one embodiment, where the network is obtainable from a composition comprising first
and second building blocks, the first building block is a polymeric molecule and the
second building block is a particle or a polymeric molecule n one embodiment, where
the network is obtainable from a composition comprising first and second building blocks,
the first building block is a polymeric molecule and the second building block is a particle.
In one embodiment, where the network is obtainable from a composition comprising a
first, the first building block is a polymeric molecule.
Polymeric Molecule
In one embodiment, a building block is a polymeric molecule.
Polymeric compounds that are covalentiy linked to cucurbiturii guest molecules are known
from VVO 2009/071 899, which is incorporated by reference herein.
Polymeric molecules comprise a plurality of repeating structural units (monomers) which
are connected by covalent bonds. Polymeric molecules may comprise a single type of
monomer (homopoiymers), or more than one type of monomer (co-polymers) . Polymeric
molecules may be straight or branched. Where the polymeric molecule is a co-polymer, it
may be a random, alternating , periodic, statistical, or block polymer, or a mixture thereof.
The co-polymer may also be a graft polymer.
In one embodiment, the polymeric molecule has 2, 3, 4 or 5 repeat units. For
convenience, such a polymer may be referred to as an oligomer.
In other embodiments, the polymeric molecule has at least 4, at least 8, at least 5, at
least 00, or at least 1,000 monomer units. The number of units may be an average
number of units.
In other embodiment, the polymeric molecule has an average number of monomer units
in a range selected from 0-200, 50-200, 50-1 50 or 75-1 25.
The number of guest molecules per polymeric molecule building block is as set out above.
Alternatively, the number of guest molecules may be expressed as the percentage of
monomers present in the polymer that are attached to guest molecules as a total of all the
monomers present in the polymeric molecule. This may be referred to as the functionality
percentage.
In one embodiment, the functionality of a polymeric molecule is at least 1 %, at least 2 %
or at least 5 %.
In one embodiment, the functionality of a polymeric molecule is at most 50 %, at most
40%, at most 20 %, at most 15 or at most 10 %.
In one embodiment, the functionality is in a range where the minimum and maximum
amounts are selected from the embodiments above. For example, the functionality is in
the range 5 to 40 %.
The functionality percentage may be determined from proton NMR measurements of a
polymer sample.
In one embodiment, the polymeric molecule has a molecular weight (Mw) of greater than
500, greater than 1000, greater than 2000, greater than 3000 or greater than 4000. The
molecular weight may be the weight average molecular weight or the number average
molecule weight. The number average and weight average molecular weights of a
polymer may be determined by conventional techniques.
In one embodiment, the polymer is a synthetic poiydisperse polymer. A polydisperse
polymer comprises polymeric molecules having a range of molecular masses. The
poiydispersity index (PD ) (weight average molecular weight divided by the number
average molecular weight) of a poiydisperse polymer is greater than , and may be in the
range 5 to 20. The po!ydispersity of a polymeric molecule may be determined by
conventional techniques such as gel permeation or size exclusion chromatography.
Suitable for use in the present invention are polymeric molecules having a relatively low
poiydispersity. Such polymeric molecules may have a polydispersity in the range
selected from 1 to 5, 1 to 3, or 1 to 2. Such polymers may be referred to as low- or
monodisperse in view of their relatively low dispersity.
The use of low- or monodisperse polymeric molecules is particularly attractive, as the
reactively of individual molecules is relatively uniform, and the products that result from
their use may also be physically and chemically relatively uniform, and may be relatively
low- or monodisperse. Methods for the preparation of low- or monodisperse polymers are
well known in the art, and include polymerisation reactions based on radical initiated
polymerisation, including RAFT (reversible addition-fragmentation chain transfer)
po!ymerisiation (see, for example, Chiefari et a . Macromolecules 1998, 31, 5559). An
example synthesis of a polymer having a low dispersity is also provided herein.
Many polymeric molecules are known in the art and may be used to produce shell
material as described herein. The choice of polymeric molecule will depend on the
particular application of the capsule. Suitable polymeric molecules include natural
polymers, such as proteins, oligopeptides, nucleic acids, glycosaminoglycans or
polysaccharides (including cellulose and related forms such as guar, chitosan chitosan,
agarose, and alginate and their functionaiised derivatives), or synthetic polymers, such as
polyethylene glycol (PEG), cis-1 ,4-polyisoprene (Pi), poly(meth)acrylate, polystyrene,
polyacrylamide, and polyvinyl alcohol. The polymer may be a homo or copolymer.
The polymeric molecule may comprise two or more natural and/or synthetic polymers.
These polymers may be arranged in a linear architecture, cyclic architecture, comb or
graft architecture, (hyper)branched architecture or star architecture.
Suitable polymeric molecules include those polymeric molecules having hydrophilic
characteristics. Thus, a part of the polymer, which part may refer to, amongst others, a
monomer unit, the backbone itself, a side chain or a grafted polymer, is hydrophilic. In
one embodiment, the polymeric molecule is capable of forming hydrogen bonds in a polar
solvent, such as water. The polymeric molecule is soluble in water to form a continuous
phase.
In one embodiment, the polymeric molecule is amphiphiiic.
Where two or more building blocks are provided, such as a first and a second building
block, each building block may be independently selected from the polymeric molecules
described above. In one embodiment, the first and second building blocks are different.
In one embodiment, the first and second building blocks are the same in this latter case,
the building blocks themselves differ only with respect to the guest molecules that are
covalently attached to each.
In one embodiment, the polymeric molecule is or comprises a poly(meth)aryclate~, a
polystyrene- and/or a poly(meth)arcylamide polymer.
In one embodiment, the polymer is or comprises a poiy(meth)aryciate polymer, which may
be or comprise a polyaryclate polymer
The acrylate functionality of the (meth)aryclate may be the site for connecting desirable
functionality, for example, for connecting a soiubiiising group or a detectable label.
In one embodiment, the polymeric molecule is obtained or obtainable from a
po yme sabie composition comprising:
(i) monomer, such as a (meth)aryciate or a styrene, which is attached to a
cucurbiturii guest molecule;
and optionally further comprising:
(ii) a monomer, such as a (meth)aryc!ate or a styrene, which is attached to a
detectable label; and/or
(iii) a monomer, such as a (meth)aryclate or a styrene, which is attached to a
soiubiiising group, such as an aquous soiubiiising group.
In one embodiment, each monomer is a (meth)aryciate monomer.
In one embodiment, each monomer is a styrene monomer.
Where (i) is present with other components, such as (ii) or (iii), it is present in the
poiymerisabie composition in at least 1, at least 5, at least 0 or at least 20 mole %.
Where (i) is present with other components, such as (ii) or (iii), it is present in the
poiymerisabie composition in at most 90, at most 50, at most 40 or at least 30 mole %.
In one embodiment, the amount of (i) present is in a range where the minimum and
maximum amounts are selected from the embodiments above. For example, the amount
present in the range 10 to 50 mole %.
In one embodiment, (i) is present at a level sufficient to provide a polymeric molecule
having a plurality of cucurbiturii guest molecules linked to each single polymer molecule.
In one embodiment, (i) is present at a level sufficient to provide a polymeric molecule
having a single cucurbiturii guest molecules linked to each single polymer molecule.
n one embodiment, (i) is present at a level sufficient to provide a polymeric molecule
having the functionality % described above.
Where (ii) is present, it is present in the poiymerisabie composition in at least 0.5, at least
1, or at least 2 mole %.
Where (ii) is present, it is present in the poiymerisab!e composition in at most 20, at most
10, or at most 5 mole %.
In one embodiment, the amount of (ii) present is n a range where the minimum and
maximum amounts are selected from the embodiments above. For example, the amount
present in the range 1 to 5 mole %.
Where (ill) s present, it is present in the po!ymerisab!e composition in at least 0.5, at least
, at least 2, at least 5, at least 0, at least 20, or at least 50 mole %
Where (ill) is present, it is present in the polymerisable composition in at most 90, at most
80, or at most 70 mole %.
In one embodiment, the amount of (iii) present is in a range where the minimum and
maximum amounts are selected from the embodiments above. For example, the amount
present in the range 10 to 80 o e %.
Where a reference is made to mole %, this is a reference to the amount of a component
present with respect to the total amount, in moles, of (i), and (ii) and (iii), where present,
and any other polymerisable monomers, where present. The component referred to may
be one of (i), (ii), (iii), or any other polymerisable monomers.
In one embodiment, the composition further comprises one or more additional
(meth)acryiate monomers. One monomer may be a (meth)acrylate monomer. One or
more monomers may be a (meth)acry!ate monomer which is substituted at the ester
group.
Where a reference is made to mole %, this is a reference to the amount of a component
present with respect to the total amount, in moles, of (i), and (ii) and (iii), where present,
and any other polymerisable monomers, where present. The component referred to may
be one of (i), (ii), (iii), or any other polymerisable monomers. The component referred to
may be a chain transfer agent or a radical initiator, as described below.
The term attached refers to the connection of the acrylate (ester), group or the phenyl
group of the styrene, either directly or indirectly to the group specified. Where there is an
indirect connected it is understood that a linker group may form the connection between
the acrylate and the group specified. In one embodiment, the linker may comprise a
(po!y)ethylene glycol (PEG) group.
In one embodiment, the detectable label is a fluorescent label. The fluorescent label may
be a fluorescein or rhodamine label. The "colour" of the label is not particularly restricted,
and green, red, yellow, cyan and orange labels are suitable for use.
In one embodiment, the aqueous soiubiiising group is a PEG group. The PEG group may
have at least 2, 3, 4, 5 or 0 repeat ethylene glycol units. The PEG group may have at
most 50, 40, 20, or 5 repeat ethylene glycol units.
In one embodiment, the aqueous solubilising group is or comprises amino, hydroxy,
carboxy, or sulfonic acid.
In one embodiment, the amino group is a quaternary amino group, for example a
trimethy!amino group.
In one embodiment, the composition further comprises a chain transfer agent.
In one embodiment, the chain transfer agent is a thiocarbony!thio compound.
Where a chain transfer agent is present, it is present in the poiymerisabie composition in
at least 0.1 , at least 0.5, or at least 1 mole %.
Where a chain transfer agent is present, it is present in the poiymerisabie composition in
at most 10, at most 5, or at most 2 mole %.
In one embodiment, the amount of a chain transfer agent present is in a range where the
minimum and maximum amounts are selected from the embodiments above. For
example, the amount present in the range 0.5 to 2 mole %.
In one embodiment, the composition further comprises a radical initiator.
Where a radical initiator is present, it is present in the poiymerisabie composition in at
least 0.01 , at least 0.05, at least 0.1 mole %.
Where a radical initiator is present, it is present in the poiymerisabie composition in at
most 5, at most 2, at most 1, or at most 0.5 mole %.
In one embodiment, the amount of a radical initiator present is in a range where the
minimum and maximum amounts are selected from the embodiments above. For
example, the amount present in the range 0.1 to 0.5 mole %.
In one embodiment, the radical initiator is selected from the group consisting of A BN
(azobisisobutyronitrile), ACPA (4,4'-azobis(4-cyanopentanoic acid)) and ACVA
(4.4'-Azobis(4-cyanova!eric acid).
In one embodiment, the polymeric molecule is obtained or obtainable from the
polymerisation of a composition comprising (i) and optionally (ii) and/or (iii) using the
change transfer agent and/or radical initiator described.
In one embodiment, the polymeric molecule is obtainable or obtained from a composition
described herein using a radical polymerisation process. In one embodiment, the
In one embodiment, the polymerisation reaction is performed at elevated temperature.
The reaction may be performed at a temperature of at least 30, at least 40 or at least
50°C.
The reaction may be performed at a temperature of at most 00, at most 90 or at most
80°C.
In one embodiment, the polymerisation reaction is performed in an organic solvent. The
original solvent may be an ether solvent, for example 1,4-dioxane, or an a ky alcohol
solvent, for example ethanoL The polymerisation reaction may be performed at reflux
temperature.
The concentration of the polymerisable mixture in the organic solvent may be at most 5.0,
at most 2.0, or at most .5 .
The concentration of the polymerisable mixture in the organic solvent may be at least
0.05, at least 0.1 , at least 0.5 M, or at least 1.0 M.
In one embodiment, the concentration is in a range where the minimum and maximum
amounts are selected from the embodiments above. For example, the concentration is in
the range 1.0 to 2.0 M.
In one embodiment, the polymerisation reaction is performed for at least , at least 5 or at
least 0 hours.
In one embodiment, the polymerisation reaction is performed for at most 72, or at most
48 hours.
The polymerisation reaction may be stopped using techniques familiar to those in the art.
Steps may include reaction mixture dilution and/or temperature reduction.
In one embodiment, the polymerisation reaction is performed for a time sufficient to obtain
a polymeric molecule having a molecular weight as described herein.
In one embodiment, the polymerisation reaction is performed for a time sufficient to obtain
a polymeric molecule having a plurality of guest molecules.
In one embodiment, the polymerisation reaction is performed for a time sufficient to obtain
a polymeric molecule having one guest molecule.
The concentration of the polymerisable mixture refers to the total amount of monomer
present (which includes (i), and (ii) and (iii). where present, and any other polymerisable
monomers, where present) in moles, in unit volume of organic solvent (i.e. per litre).
In one embodiment, the polymer may be formed as a particle.
Particle
In one embodiment, the building block is a particle. The type of particle for use in the
present invention is not particularly limited.
In one embodiment, the particle is a first building block and the particle is linked to a
plurality of cucurbituril guest molecules.
In one embodiment, the particle is a second building block and the particle is linked to one
or more cucurbituril guest molecules.
In one embodimeni, the particle is a second building block and the particle is linked to a
plurality of cucurbituril guest molecules.
Typically, the particle has a size that is one, two, three or four magnitudes smaller than
the size of the capsule.
In one embodiment, the particle is a nanoparticle. A nanoparticle has an average size of
at least 1, at least 5, or at least 0 nm in diameter. A nanoparticle has an average size of
at most 900, at most 500, at most 200, or at most 00 nm in diameter.
In one embodiment, the nanoparticle has an average size in the range 1-100 nm or
5-80 nm in diameter.
The average refers to the numerical average. The diameter of a particle may be
measured using microscopic techniques, including TE .
In one embodiment, the particles have a relative standard deviation (RSD) of at most
0.5%, at most 1%, at most .5%, at most 2%, at most 4%, at most 5%, at most 7%, at
most 10%, at moist 15 %, at most 20 % or at most 25 %.
In one embodiment, the particle has a hydrodynamic diameter of at least 1, at least 5, or
at least 10 nM in diameter.
In one embodiment, the particle has a hydrodynamic diameter of at most 900, at most
500, at most 200, or at most 100 nM in diameter.
The hydrodynamic diameter may refer to the number average or volume average. The
hydrodynamic diameter may be determined from dynamic light scattering (DLS)
measurements of a particle sample.
In one embodiment, the particle is a metal particle.
In one embodiment, the particle is a transition metal particle.
In one embodiment, the particle is a noble metal particle.
In one embodiment, the particle is or comprises copper, ruthenium, palladium, platinum,
titanium, zinc oxide, gold or silver, or mixtures thereof.
In one embodiment, the particle is or comprises gold, silver particle, or a mixture thereof.
In one embodiment, the particle is a gold or a silver particle, or a mixture thereof.
In one embodiment, the particle is a gold nanoparticle (AuNP).
In one embodiment, the particle is or comprises silica or calcium carbonate.
In one embodiment, the particle is a quantum dot.
In one embodiment, the particle is or comprises a polymer. The polymer may be a
polystyrene or poiyacrylamide polymer. The polymer may be a biological polymer
including for example a polypeptide or a polynucleotide.
In one embodiment, the particle comprises a material suitable for use in surface
enhanced Raman spectroscopy (SERS). Particles of gold and/or silver and/or other
transition metals are suitable for such use.
Gold and silver particles may be prepared using techniques known in the art. Examples
of preparations include those described by Coulston et al. (Chern. Commun. 20 , 47,
164) Martin et al. (Martin et al. Langmuir 2010, 26, 74 0) and Frens (Frens Nature Phys.
Sci. 1973, 241, 20), which are incorporated herein by reference in their entirety.
The particle is linked to one or more guest molecules, as appropriate. Typically, where
the particle is a first building block, it is provided at least with a plurality of guest
molecules. Where, the particle is a second building block, it is provided at one or more
guest molecules.
In one embodiment, a guest molecule ma be covalentiy linked to a particle via a linking
group. The linking group may be a spacer element to provide distance between the guest
molecule and the particle bulk. The linker may include functionality for enhancing the
water solubility of the combined building block and guest molecule construct. The linker is
provided with functionality to allow connection to the particle surface. For example, where
the particle is a gold particle, the linker has thiol functionality for the formation of a
connecting gold-sulfur bond.
Alternatively, a guest molecule may be attached directly to the particle surface, through
suitable functionality. For example, where the particle is a gold particle, the guest
molecule may be attached to the gold surface via a thiol functionality of the guest
molecule.
In one embodiment, the particle comprises solubilising groups such that the particle,
together with its guest molecules, is soluble in water or is soluble in a water immiscible
phase.
The solubilising groups are attached to the surface of the particle. The solubilising group
may be covalentiy attached to the particle through suitable functionality. Where the
particle is a gold particle, the solubilising group is attached through a sulfur bond to the
gold surface.
The solubilising group may be, or comprise, polyethylene glycol or amine, hydroxy,
carboxy or thiol functionality.
In one embodiment, the building block is obtained or obtainable from a composition
comprising:
(i) a gold particle;
(ii) a guest molecule together with a linking group that has thiol functionality; and
(iii) a soiubilising molecule having thiol functionality; and optionally further
comprising
(iv) a further guest molecule, together with a linking group that has thiol
functionality.
In one embodiment, the amount of guest molecule present in the composition is at least 1,
at least 5, at least 0 or at least 5 mole %.
In one embodiment, the amount of guest molecule present in the composition is at most
80, at most 50, or most 25 mole %.
A reference to mole % is a reference to the amount of guest molecule present as a
percentage of the total amount of (ii) and (iii), and (iv) where present, in the composition.
The amount of (ii) present in the composition may be such to allow the preparation of a
particle building block having a plurality of guest molecules.
Cucurbituri! Guest
As noted above, the guest is a compound that is capable of forming a guest-host complex
with a cucurbiturii. The term compiexation therefore refers to the establishment of the
guest-host complex.
In some embodiments of the invention, the guest host complex is a ternary complex
comprising the cucurbiturii host and a first guest molecule and a second molecule.
Typically such complexes are based around CB[8] and variants and derivatives thereof.
In some embodiments of the invention, the guest host complex is a binary complex
comprising the cucurbiturii host and a first guest molecule. Typically such complexes are
based around CB[5] or CB[7], and variants and derivatives thereof. In the present
invention, binary complexes are obtainable from a plurality of covalently linked
cucurbiturils. CB[8], and variants and derivatives thereof, may also form binary
complexes.
In principal, any compound having a suitable binding affinity may be used in the methods
of the present invention. The compound used may be selected based on the size of the
moieties that are thought to interact with the cavity of the cucurbiturii. The size of these
moieties may be sufficiently large to permit compiexation only with larger cucurbiturii
forms.
The term selective may be used to refer to the amount of guest-host complex formed.
where the cucurbiturii (the first cucurbiturii) and a second cucurbiturii are present in a
mixture with a particular guest molecule or guest molecules. The guest-host complex
formed between the first cucurbiturii and the guest (in a binary compiex) or guests (in a
ternary complex) may be at least 60 mo %, at least 70 mo! %, at least 80 mol %, at least
90 mol %, at least 95 mol %, at least 97 mol %, at least 98 mol %, or at least 99 mol %, of
the total amount of guest-host complex formed (for, example taking into account the
amount of guest-host complex formed between the second cucurbiturii and the guest or
guests, if any).
In one embodiment, the guesi-host complex formed from the (first) cucurbiturii and the
guest or guests has a binding affinity that is at least 00 times greater than the binding
affinity of a guest host complex formed from the second cucurbiturii and the guest or
guests. Preferably, the binding affinity is at least 103, at least 104, at least 105, at least
106, or at least 10 greater.
Cucurbiturii guest molecules are well known in the art. Examples of guest compounds for
use include those described in VVO 2009/071 899, Jiao et a . (Jiao et a . Org. Lett 201 1,
13, 3044), Jiao et ai. (Jiao et ai. J. Am. Chem. Soc 2010, 132, 15734) and Rauwaid et aL
(Rauwald et al. J. Phys. Chem. 2010, 114, 8806).
Described below are guest molecules that are suitable for use in the formation of a
capsule shell. Such guest molecules may be connected to a building block using
standard synthetic techniques.
A cucurbiturii guest molecule may be derived from, or contain, a structure from the table
below:

where the structure may be a salt, including protonated forms, where appropriate. In one
embodiment, the guest moiecuies are guest molecules for CB[8].
In one embodiment, the guest molecule is, or is derived from, or contains, structure
A1-A43, A46 or B 1-B4, in the table above.
In one embodiment, the guest molecule is, or is derived from, or contains, structure A ,
A2, or A 3 in the table above.
In one embodiment, the guest molecule is, or is derived from, or contains, structure B 1.
Additionally, the guest molecule is or is derived from, or contains, adamantane, ferrocene
or cyclooctane (including bicyclo[2.2.2]octane). Such are described by Moghaddam et a .
(see J. Am. Ch . Soc. 201 1, 133, 3570).
In some embodiments, first and second guest molecules form a pair which may interact
within the cavity of cucurbiturii to form a stable ternary host-guest complex. Any guest
pair that fits within the cavity of the cucurbiturii may be employed. In some embodiments,
the pair of guest molecules may form a charge transfer pair comprising an electron-rich
and an electron-deficient compound. One of the first and second guest moiecuies acts as
an electron acceptor and the other as an electron donor in the CT pair. For example, the
first guest molecule may be an electron deficient molecule which acts an electron
acceptor and the second guest molecule may be an electron rich molecule which acts as
an electron donor or vice versa. In one embodiment, the cucurbituri! Is CB[8].
Suitable electron acceptors include 4,4'-bipyridinium derivatives, for example
. V dimetby dipy dy iumyiethy ene, and other related acceptors, such as those based on
diazapyrenes and diazaphenanthrenes. Vioiogen compounds including a ky vio!ogens
are particularly suitable for use in the present invention. Examples of a kyl vioiogen
compounds include A/,/S/'-dimethyl-4,4'-bipyridinium salts (also known as Paraquat)
Suitable electron donors include electron-rich aromatic molecules, for example ,2-
dihydroxybenzene, 1,3-dihydroxybenzene, 1,4-dibydroxybenzene, tetrathiafulvalene,
naphthalenes such as 2,6-dihydroxynaphthaiene and 2-naphthol, indoles and sesamo!
(3,4-methyienedioxyphenol). Poiycyclic aromatic compounds in general may find use as
suitable electron donors in the present invention. Examples of such compounds include
anthracene and naphthacene
Amino acids, such as tryptophan, tyrosine and phenylalanine may be suitable for use as
electron donors. Peptide sequences comprising these amino acids at their terminus may
be used. For example, a donor comprising an amino acid sequence N-VVGG-C,
N-GGW-C or N-GWG-C may be used.
In some embodiments, the guest molecules are a pair of compounds, for example first
and second guest molecules, where one of the pair is an A compound as set out in the
table above (e.g. A , A2, A3 etc), and the other of the pair is a B compound as set out in
the table above (e.g. B 1 , B2, B3 etc.). In one embodiment, the A compound is selected
from A -A43 and A46. In one embdoiemtn, the B compound is B .
Other suitable guest molecules include peptides such as VVGG (Bush, M. E. et al J. Am.
Che Soc. 2005, 127, 1451 1-1451 7).
An electron-rich guest molecule may be paired up with any electron-deficient CB[8] guest
molecule. Examples of suitable pairs of guest molecules for example first and second
guest molecules, for use as described herein may include:
vioiogen and naphtho!;
vioiogen and dihydroxybenzene;
vioiogen and tetrathiafulvalene;
vioiogen and indole;
methyivioiogen and naphthoi;
methylvioiogen and dihydroxybenzene;
methyivioiogen and tetrathiafulvalene;
methyivioiogen and indole;
/,/V-dimethyidipyridyiiumylethyiene and naphthoi;
/V,/V-dimethy!dipyridy!iumylethy!ene and dihydroxybenzene;
A/,A/-dimethyidipyridyiiumyleihy!ene and tetrathiafulvalene;
/S/,/V-dimeihy!dipyridy!iumylethyiene and indole;
2,7-dimethyldiazapyrenium and naphtho!;
2,7-dimethyldiazapyrenium and dihydroxybenzene;
2,7-dimethyldiazapyrenium and tetrathiafulvalene; and
2,7-dimethyldiazapyrenium and indole.
In particular, suitable pairs of guest molecules for use as described herein may include
2-naphtho! and methyl viologen, 2,6-dihydroxynaphtha!ene and methyl viologen and
tetrathiafulvalene and methyl viologen.
In one embodiment, the guest pair is 2-naphtho! and methyl viologen.
in one embodiment, the guest pair is a reference to a pair of guest molecules suitable for
forming a ternary complex with CB[8].
in one embodiment, the guest molecule is preferably an ionic liquid. Typically, such
guests are suitable for forming a complex with CB[7]. However, they may also form
complexes with CB[8] in either a binary complex, or in a ternary complex together with
another small guest molecule or solvent (see Jiao et a . Org. Lett. 20 , 13, 3044).
The ionic liquid typically comprises a cationic organic nitrogen heterocycie, which may be
an aromatic nitrogen heterocycie (a heteroaryl) or a non aromatic nitrogen heterocycie.
The ionic liquid also typically comprises a counter-anion to the cationic organic nitrogen
heterocycie. The nitrogen heteroaryl group is preferably a nitrogen C -ioheteroary! group,
most preferably a nitrogen C 6heteroary! group, where the subscript refers to the total
number of atoms in the ring or rings, including carbon and nitrogen atoms. The non
aromatic nitrogen heterocycie is preferably a nitrogen C heterocyeie, where the subscript
refers to the total number of atoms in the ring or rings, including carbon and nitrogen
atoms. A nitrogen atom in the ring of the nitrogen heterocycie is quaternised.
The counter-anion may be a halide, preferably a bromide. Other counter-anions suitable
for use are those that result in a complex that is soluble in water.
The guest is preferably a compound, including a salt, comprising one of the following
groups selected from the list consisting of: irnidazo!ium moiety; pyridinium moiety;
quino!inium moiety; pyrimidinium moiety; pyrrolium moiety; and quaternary pyrrolidine
moiety.
Preferably, the guest comprises an imidazo!ium moiety. An especially preferred guest is
1-aikyi-3-alkylimidazolium, where the a ky groups are optionally substituted.
1-Alkyl-3-aikyiimidazoiium compounds, where the a ky groups are unsubstituted, are
especially suitable for forming a complex with CB[7].
1-Alkyl-3-aikyiimidazoiium compounds, where the alkyl groups are unsubstituted, are
especially suitable for forming a complex with CB[8]
1-Alkyl-3-aikyiimidazoiium compounds, where an alkyl group is substituted with aryl
(preferably napthyl), are especially suitable for forming a complex with CB[8].
The -alkyl and 3-a ky substituents may the same or different. Preferably, they are
different.
n one embodiment, the 3-alkyl substituent is methyl, and is preferably unsubstituted.
In one embodiment, the -alkyl substituent is ethyl or butyl, and each is preferably
unsubstituted.
In one embodiment, the optional substituent is aryl, preferably C ..ioaryi. Aryl includes
carboaryi and heteroaryi. Aryl groups include phenyl, napthyl and quinoiinyl.
In one embodiment, the alkyl groups described herein are linear alkyl groups.
Each alkyl group is independently a - a!ky! group, preferably a C alky! group.
The aryl substituent may itself be another 1-a!ky!-3-substituted-imidazolium moiety (where
the alkyl group is attached to the 3-position of the ring).
In another embodiment, the compound preferably comprises a pyridinium moiety.
The ionic liquid molecules describe above are particular useful for forming binary guesthost
complexes. Complexes comprising two ionic liquid molecules as guests within a
cucurbiturii host are also encompassed by the present invention.
A cucurbiturii may be capable of forming both binary and ternary complexes. For
example, it has been previously noted that CB[6] compounds form ternary complexes with
short chain 1-a!ky!-3-methylimidazolium guest molecules, whilst longer chain 1-aSkyl-3-
methyiimidazoiium guest molecules form binary complexes with the cucurbiturii host.
Preferred guests for use in the present invention are of the form H+X , where H+ is one of
the following cations,
and X is a suitable counter-anion, as defined above. A preferred counter anion is
a halide anion, preferably Br.
In a preferred embodiment cation A or cation B may be used to form a complex with
CB[7] or CB[6].
In a preferred embodiment, cation D or cation E may be used to form a complex with
CB[8]
Cations A and B may be referred to as 1-ethyI-3-methy!imidazo!ium and 1-butyI-3-
methy!imidazo!ium respectively.
Cations D and E may be referred to as 1-naphthaieny!methyl-3-methy!imidazo!ium, where
D is 1-naphthalen-2-ylmethyl-3-methyiimidazoiium and E is 1-naphtha!en-1-ylmethyi-3-
methyiimidazoiium.
Alternatively or additionally, the guest compounds may be an imidazolium salt of formula
(I):
"
wherein X is a counter anion;
R is independently selected from H and saturated C - a ky ;
R2 is independently C alkyl which may optionally contain one or more double or
triple bonds, and may be optionally interrupted by a heteroatom selected from -0-, -S-,
-NH-, and -B-, and may be optionally substituted.
In one embodiment, X is independently selected from the group consisting of C , Br , G,
BF,, , PF6 , OH , 8 H , HBO/, HC0 3 , NTf , C N50 , AICi , Fe3Ci N0 3 , N e8 , eS0 3 ,
SbFe , PrCB H·· . AuC , HF2 , N0 2 , Ag(CN)2 , and NiC . In one embodiment, X is
selected from C , Br , and G.
In one embodiment, R is selected from H and linear saturated C - alkyl.
In one embodiment, R2 is linear C alkyl, which may optionally contain one or more
double bonds, and may be optionally interrupted by a heteroatom selected from -0-, -S-,
-NH-, and -B-, and may be optionally substituted.
In one embodiment, R2 is linear C,-1 alkyl, which may optionally contain one or more
double bonds, and may be optionally substituted.
In one embodiment, where a double or triple bond is present, it may be conjugated to the
imidazolium moiety. Alternatively, the double or triple bond may not be conjugated to the
imidazolium moiety.
In one embodiment, the optional substituents are independently selected from the group
consisting of halo, optionally substituted C -2oaryl, -OR3, -OCOR3, =0, -SR , =S, -BR3, -
R3R4 -NR3COR 3, -N(R )CONR R4, -COOR 3, -C(0)R 3, -C(=0)SR 3, -CONR R4, -G(8)R 3,
-C(=S)SR 3, and -C(=S)NR R4,
where each of R3 and R4 is independently selected from H and optionally
substituted saturated - aiky , C5-20 y and C -6 a!ky!ene-C -2o l
or R3 and R4 may together may form an optionally saturated 5-, 6- or 7-membered
heterocyclic ring which is optionally substituted with a group -R3.
In one embodiment, the optional substituents are independently selected from the group
consisting of halo, optionally substituted C5-20 aryl, -OR3, -QCOR 3, -NR R4. -NR COR 3,
-N(R 3)CONR 3R4, -COOR 3, -C(0)R 3, and -CONR R4, where R3 and R4 are defined as
above.
Each C - 0 aryl group may be independently selected from a Ce-20 carboaryi group or a C .
20 heteroaryi group.
Examples of - carboaryi groups include phenyl and napthyi.
Examples of C5-20 heteroaryi groups include pyrrole (azo!e) (C ) , pyridine (azine) (C6) ,
furan (oxoie) (C5) , thiophene (thioie) (C5) , oxazole (C ) , thiazole (C5) , imidazole
( ,3-diazoie) (C5) , pyrazole ( ,2-diazoie) (C5) , pyridazine ( ,2-diazine) (C6) , and pyrimidine
( ,3-diazine) (C6) (e.g., cytosine, thymine, uracil).
Each C5-20 aryl is preferably selected from optionally substituted phenyl, napthyi and
imidazolium.
Each C5-20 aryl group is optionally substituted. The optional substituents are
independently selected from halo, C1-6 aiky!, -OR3, -OCOR 3, -NR R4, -NR COR3,
-N(R )CONR R4, -COOR 3, -C(0)R 3, and -CONR R4, where R3 and R4 are defined as
above.
In one embodiment, each C .2 o aryl group is optionally substituted with C 6 a ky .
Where the C o aryl group is an imidazolium, such is preferably substituted at nitrogen
with a group R (thereby forming a quaternary nitrogen).
The compound of formula (I) comprises an imidazolium moiety having a substituent R2 at
the 1-position and a substituent R at the 3-position. In a further aspect of the invention,
the compound of formula (I) may be optionally further substituted at the 2-, 4- or 5-positon
with a group R , wherein R has the same meaning as R .
The embodiments above are combinabie in any combination, as appropriate.
Encapsulant
The capsule of the invention may be used to encapsulate a component (the encapsulant).
In one embodiment there is provided a capsule comprising an encapsulant. The capsule
is suitable for storing a component, and this component may be later released as required
at a chosen location.
It is understood that a reference to an encapsulated component is not a reference to a
solvent molecule. For example, the encapsulated component is not water or is not an oil
or an organic solvent. It is also understood that a reference to an encapsulated
component is not a reference to a cucurbiturii or a building block for use in the preparation
of the capsule shell. Otherwise, the component is not particularly limited.
The encapsulant is therefore a component of the capsule that is provided in addition to
solvent that may be present within the shell.
In the methods of the invention the capsule shell is prepared from a composition
comprising a cucurbiturii and one or more building blocks, as appropriate. Not all the
cucurbiturii and one or more building blocks may react to form shell material. Additionally,
the cucurbiturii and one or more building blocks may react to form a network, but this
network may be not be included in the shell that forms the capsule. These unreacted or
partially reacted reagents and products may be contained within the shell, and may be
contained in addition to the encapsulant. Thus, the encapsulant is a component of the
capsule that is provided in addition to unreacted or partially reacted reagents and
products that may be present within the shell.
In one embodiment, the encapsulant compound has a solubility of at least 0.01 mg/mL, at
least 0.02 mg/mL, at least 0.05 mg/mL, or at least 0.10 mg/mL.
In one embodiment, the solubility refers to aqueous solubility (i.e. an aqueous phase).
In one embodiment, the solubility refers to solubility in an oil phase or an organic phase.
The capsules of the invention may be used to encapsulate a wide range of components.
In one embodiment, the encapsulated component has a molecular weight of at least 100,
at least 200, at least 300, at least 1,000, at least 5,000 ( 1 k), at least 10,000 (10k), at least
50,000 (50k), at least 100,000 ( 100k) or at least 200,000 (200k).
In one embodiment, the encapsulant is a therapeutic compound.
In one embodiment, the encapsulant is a biological molecule, such as a polynucleotide
(for example DNA and RNA), a polypeptide or a polysaccharide.
In one embodiment, the encapsulant is a polymeric molecule, including biological
polymers such as those polymers mentioned above.
In one embodiment, the encapsuiant is a cell
in one embodiment, the encapsuiant is an ink.
In one embodiment, the encapsuiant is a carbon nanotube.
In one embodiment, the encapsuiant is a particie. The particle may be a metal particle.
The size of the capsule is selected so as to accommodate the size of the encapsuiant.
Thus, the internal diameter (the distance from innermost wall to innermost wail) is greater
than the greatest dimension of the encapsuiant.
n one embodiment, the encapsuiant has a detectable label. The detectable label may be
used to quantify and/or locate the encapsuiant. The label may be used to determine the
amount of encapsuiant contained with the capsule.
In one embodiment, the detectable label is a luminescent label. In one embodiment, the
detectable label is a fluorescent label or a phosphorescent label.
In one embodiment, the detectable label is a visible.
In one embodiment, the fluorescent label is a rhodamine or fluorescein label.
n one embodiment, the capsule of the invention is for use as a reactor. The method of
preparing the capsule as described herein brings together the reagents, which are
supplied in separate second phase sub-flows, and are contacted at substantially the same
time as the second phases contact the first phase. A shell of material is formed at the
interface of the discrete regions that is formed, and this shell contains the reagents which
may be permitted to react. The localisation of reagents within a discrete region is allows
control over reaction timings.
Where the capsule is for use as a microreactor it is understood that the composition of the
shell inner space will change over time as the reagents react to form a product, along with
associated by-products, if any. As will be apparent, the amount of reagent wil decrease
as the reaction progresses.
Additional and Alternative Encapsulants
In addition to, or as alternatives to, the encapsulants mentioned above, the encapsuiant
may be selected from one or more of the encapsulants discussed below. In one
embodiment, the molecular weight preferences given above apply to these encapsulants.
In one embodiment, the encapsuiant is selected from the group consisting of toxic
molecules (such as nerve agents and heavy metals), hormones, herbicides, pesticides,
antibodies, pathogens (such as viruses), adjuvants, gels, nanoparticles (including metal or
non-metal particles), polymers (including synthetic and natural polymers), catalysts
(organic, inorganic, and organometal!ic), adhesives and sealants.
A pathogen is an agent that is capable of causing disease in a host. The pathogen may
be a virus, a bacterium, a fungus, or a prion.
In one embodiment, the encapsulant is a virus.
The virus may be virus selected from a family selected from the group consisting of
adenoviridae (e.g. adenovirus), herpesviridae (e.g. Herpes simplex, type 1 and type 2,
and Epstein-barr), papiliomaviridae (e.g. human papillomavirus), hepadnaviridae (e.g.
Hepatitis B), flaviviridae (e.g. Hepatitis C, yellow fever, dengue, West Nile), retroviridae
(e.g. immunodeficiency virus (HIV)), orthomyxoviridae (e.g. Influenza), paramyxoviridae
(e.g. measles, mumps), rhabdoviridae (e.g. rabies), and reoviridae (e.g. rotavirus).
In one embodiment, the encapsulant is a microorganism
As noted above, in one embodiment, the encapsulant is a ceil. The cell may be a
prokaryotic or a eukaryotic ceil.
The cell may be a mammal cell, such as a human cell, a rodent ceil (e.g., a guinea pig, a
hamster, a rat, a mouse) a iagomorph ceil (e.g., a rabbit), an avian cell (e.g., a bird), a
canine cell (e.g., a dog), a feline cell (e.g., a cat), an equine ceil (e.g., a horse), a porcine
cell (e.g., a pig), an ovine cell (e.g., a sheep), a bovine cell (e.g., a cow), a simian cell
(e.g., a monkey or ape), a monkey ce l (e.g., marmoset, baboon), an ape cell (e.g., gorilla,
chimpanzee, orangutang, gibbon), or an ornithorhynchidae ceil (e.g. platypus).
The cell may be a tumour ceil, which may be a benign or malignant tumour cell.
Examples of eukaryotic ceils include epithelial, endotherial, neural, skeletal, and fibroblast
cells, amongst others.
In one embodiment, the encapsulant is a bacterium, such as a gram positive bacterium
and a gram negative bacterium.
Examples of gram positive bacteria include Corynebacierium, Mycobacterium, Nocardia,
Streptomyces, Staphylococcus (such as S. aureus), Streptococcus (such as
S. pneumoniae), Enterococcus (such as E. faecium), Bacillus, Clostridium (such as C.
diff.) and Listeria.
Examples of gram negative bacteria include Hemophilus, Klebsiella, Legionella,
Pseudonnonas, Escherichia (such as E. coii), Proteus, Enterobacter, Serratia,
Helicobacter {such as Helicobacter pylon), and Salmonella.
in one embodiment, the encapsulant is an antibody.
The term "antibody" herein is used in the broadest sense and specifically covers
monoclonal antibodies, polyclonal antibodies, dimers, muitimers, muitispecific antibodies
{e.g., bispecific antibodies), and antibody fragments, so long as they exhibit the desired
biological activity. Antibodies may be murine, human, humanized, chimeric, or derived
from other species. An antibody is a protein generated by the immune system that is
capable of recognizing and binding to a specific antigen. A target antigen generally has
numerous binding sites, also called epitopes, recognized by CDRs on multiple antibodies.
Each antibody that specifically binds to a different epitope has a different structure. Thus,
one antigen may have more than one corresponding antibody. An antibody includes a fulllength
immunoglobulin molecule or an immunologically active portion of a full-length
immunoglobulin molecule, i.e., a molecule that contains an antigen binding site that
immunospecifically binds an antigen of a target of interest or part thereof, such targets
including but not limited to, cancer cell or cells that produce autoimmune antibodies
associated with an autoimmune disease. The immunoglobulin can be of any type (e.g.
gG, gE, igM, IgD, and IgA), class (e.g. lgG1 , igG2, lgG3, lgG4, igA1 and lgA2) or
subclass of immunoglobulin molecule. The immunoglobulins can be derived from an
species, including human, murine, or rabbit origin.
"Antibody fragments" comprise a portion of a full length antibody, generally the antigen
binding or variable region thereof. Examples of antibody fragments include Fab, Fab',
F(ab')2, and Fv fragments; diabodies; linear antibodies; fragments produced by a Fab
expression library, anti-idiotypic (anti-Id) antibodies, CDR (complementary determining
region), and epitope-binding fragments of any of the above which immunospecifically bind
to cancer cell antigens, viral antigens or microbial antigens, single-chain antibody
molecules; and mu!tispecific antibodies formed from antibody fragments.
The term "monoclonal antibody" as used herein refers to an antibody obtained from a
population of substantially homogeneous antibodies, i.e. the individual antibodies
comprising the population are identical except for possible naturally occurring mutations
that may be present in minor amounts. Monoclonal antibodies are highly specific, being
directed against a single antigenic site. Furthermore, in contrast to polyclonal antibody
preparations which include different antibodies directed against different determinants
(epitopes), each monocional antibody is directed against a single determinant on the
antigen. In addition to their specificity, the monocional antibodies are advantageous in
that they may be synthesized uncontaminated by other antibodies. The modifier
"monoclonal" indicates the character of the antibody as being obtained from a
substantially homogeneous population of antibodies, and is not to be construed as
requiring production of the antibody by any particular method. For example, the
monoclonal antibodies to be used in accordance with the present invention may be made
by the hybridoma method, or may be made b recombinant DNA methods. The
monoclonal antibodies may also be isolated from phage antibody libraries.
The monoclonal antibodies herein specifically include "chimeric" antibodies in which a
portion of the heavy and/or light chain is identical with or homologous to corresponding
sequences in antibodies derived from a particular species or belonging to a particular
antibody class or subclass, while the remainder of the chain(s) is identical with or
homologous to corresponding sequences in antibodies derived from another species or
belonging to another antibody class or subclass, as well as fragments of such antibodies,
so long as they exhibit the desired biological activity
In one embodiment, the antibody is an antibody-drug conjugate (ADC).
The antibody may be suitably labelled for detection and analysis, either whilst held in the
capsule, of for later use, when the antibody is released.
In one embodiment, the encapsuiant is a hormone. The hormone may a peptidic
hormone, such as insulin or growth hormone, or a lipid hormone, such as a steroid
hormone, for example prostaglandin and estrogen.
In one embodiment, the encapsuiant is a polypeptide. In one embodiment, the
polypeptide is a protein in one embodiment the protein has catalytic activity, for example
having iigase, isomerase, lyase, hydrolase, transferase or oxidoreductase activity.
In one embodiment, the encapsuiant is a polymer. In some embodiments, the capsule
shell of the present invention includes a building block that is a functionalised polymer.
Where such a building block is present, a polymer that is an encapsuiant differs from the
building block. In one embodiment, the encapsuiant polymer is not suitable for forming a
non-cova!ent link with a cucurbituri!.
In one embodiment, the encapsuiant is a metal particle.
In one embodiment, the nanoparticle is or comprises a noble metal.
In one embodiment, the nanoparticle is or comprises a transition metal.
In some embodiments, the capsule shell of the present invention includes a building block
that is a functionalised particle. Where such a building block is present, a particle that is
an encapsuiant differs from the building block. In one embodiment, the encapsuiant
particle is not suitable for forming a non-covalent link with a cucurbituri!.
In one embodiment, the nanoparticle is a gold nanoparticle (AuNP) or a silver
nanoparticle (AgNP), or a nanoparticle comprising both silver and gold.
Generally, the particle is substantially spherical. However, particles having other shapes
may be used, if appropriate or desirable.
In one embodiment, the nanoparticle has a diameter of at most 500 nm, at most 200 nm,
at most 150 nm, at most 100 nm, at most 80 nm, or at most 70 nm.
In one embodiment, the nanoparticle has a diameter of at least 1 nm, at least 2 nm, at
least 5 nm, at least 0 nm, at least 15 nm, at least 20 nm, at least 30 nm, or at least
40 nm.
In one embodiment, the diameter of the particle is in a range where the minimum and
maximum rates are selected from the embodiments above. For example, the diameter is
in the range 1 to 100 nm, or for example in the range 10 to 100 nm. For example, the
diameter is in the range 2 to 500 nm
In one embodiment, the nanoparticle has a diameter of about 20 nm.
The average refers to the numerical average. The diameter of a particle may be
measured using microscopic techniques, including TE .
The particles used in the present invention are sustainab!y monodisperse or have a very
low dispersity. In one embodiment, the particles have a relative standard deviation (RSD)
of at most 0.5%, at most 1%, at most 1.5%, at most 2%, at most 4%, at most 5%, at most
7%, at most 10%, at moist 5 %, at most 20 % or at most 25 %.
In one embodiment, the particle has a hydrodynamic diameter of at least 5 nm, at least 10
nm, at least 15 nm, at least 20 nm, at least 30 nm, at least 40 nm.
In one embodiment, the particle has a hydrodynamic diameter of at most 500 nm, at most
200 nm, at most 150 nm, at most 100 nm, at most 80 nm, or at most 70 nm.
The hydrodynamic diameter may refer to the number average or volume average. The
hydrodynamic diameter may be determined from dynamic light scattering (DLS)
measurements of a particle sample.
The size of the particle and the composition of the particle may be selected to provide the
most appropriate or beneficial surface enhanced effect.
Gold and silver particles may be prepared using techniques known in the art. Examples
of preparations include those described by Coulston (Couiston et a Chem. Commun.
201 , 47, 164} and Martin (Martin et al. Langmuir 2010, 26 7410) and Frens (Frens
Nature Phys. Sci 1973, 241, 20), which are incorporated herein by reference in their
entirety.
In one embodiment, the encapsulant is a polymer. In one embodiment, the polymer is not
a polymer that is present as building block in the capsule shell. Otherwise, the polymer is
not particularly limited.
Further Additional and Alternative Encapsuiants
In addition to, or as alternatives to, the encapsuiants mentioned above and the additional
and alternative encapsuiants mentioned above, the encapsuiant may be selected from
one or more of the further encapsuiants discussed below. In one embodiment, the
molecular weight preferences given above apply to these encapsuiants.
In one embodiment, the encapsuiant is a fragrance compound or a fragrance
composition. A fragrance compound or composition has suitable odorant properties for
use in a perfume.
In one embodiment, the encapsuiant s a fiavourant compound or a flavourant
composition. A flavourant may be or include a flavour enhancer, such as a sweetener.
In one embodiment, the encapsuiant is an oil, such as an essential oil. Examples of
essential oils include those obtained or obtainable from sweet orange, peppermint, lemon
and clove, amongst others.
In one embodiment, the encapsuiant is itself a vehicle for holding a encapsuiant within.
For example, the encapsuiant may be a liposome, micelle, or vesicle. The liposome,
micelle, or vesicle may hold an encapsuiant, such as one of the encapsuiants described
herein. Suitably loaded liposomes, micelles, or vesicles may be prepared using standard
techniques known in the art. The loaded liposome, micelle, or vesicle may then be
encapsulated into the supramo!ecuiar capsules of the invention using the methods
described herein.
Methods for the Preparation of Capsules
In a second aspect of the invention there is provided a method for the preparation of a
capsule having a shell, such as the capsule of the first aspect of the invention, the method
comprising the steps of:
(i) contacting a flow of a first phase and a flow of a second phase in a channel,
thereby to generate in the channel a dispersion of discrete regions, preferably droplets, of
the second phase in the first phase, wherein the second phase comprises cucurbituril and
one or more building blocks having suitable cucurbituril guest functionality suitable to form
a supramoleeular cross-linked network, thereby to form a capsule shell within the discrete
region, wherein the first and second phases are immiscible.
In one embodiment, the second phase comprises either (a) a cucurbituril and ( 1 ) or (2); or
(b) a plurality of covalentiy linked cucurbituri!s and ( 1 ) , (2) or (3), thereby to form a
capsule shell within the discrete region, wherein the first and second phases are
immiscible.
In one embodiment, the second phase comprises a cucurbituril and ( 1 ) or (2).
In one embodiment, the second phase comprises a cucurbituril and ( ) .
In the method of the invention a dispersion of the second phase is created within the
continuous first phase. In one embodiment, one of the first and second phases is an
aqueous phase and the other phase is a water immiscible phase.
In one embodiment, the second phase is an aqueous phase. The first phase is a water
immiscible phase, for example an oil phase.
In one embodiment, the first phase is an aqueous phase. The second phase is a water
immiscible phase, for example an oil phase.
In one embodiment, the method further comprises the step of (ii) collecting the outflow
from the channel, thereby to obtain a droplet, within which is a capsule.
In one embodiment, the method comprises the step (ii) above and (iii) optionally drying
the capsule obtained in step (ii). The drying step refers to the desolvation of the droplet
and the capsule. Where the second phase is an aqueous phase, the drying step is a
dehydration.
In one embodiment, the flow of the second (dispersed) phase is a flow generated by the
combination of a plurality of sub-flows, where each sub-flow comprises a reagent for use
in the preparation of the shell. Thus, one sub flow may comprise a cucurbituril (as in case
(a)) or a plurality of covaientiy linked cucurbiturils (as in case (b)). Further sub-flows may
comprise a first building block and a second building block (as with composition (1)). The
first and second building blocks may be contained within the same or different sub-flows.
In one embodiment cucurbituril and the building block (or blocks) are provided in separate
sub-flows.
In one embodiment, a sub-flow is provided for each reagent for use in the preparation of
the shell. In this embodiment, it is possible to independently alter the flow rate of each
sub-flow, thereby independently altering the final concentration of a particular reagent in
the formed discrete region. The ability to independently alter the flow rate and therefore
reagent concentration allows control over the structure of the shell formed. Thus, the
pore size and the thickness of the shell may be controlled by appropriate changes in the
sub- flow rates.
The sub-flows may be brought into contact prior to contact with the flow of the first phase.
In this arrangement, multiple sub-flows may be brought into contact at the same time, or
may be contacted in a sequence. Alternatively, the sub-flows may be brought into contact
at substantially the same time as the second phases are brought into contact with the flow
of the first phase.
In order to minimise the formation of an unstructured aggregation, including for example a
hydrogel, within the second phase, the cucurbituril or the plurality of covalently linked
cucurbiturils are brought into contact with the building blocks immediately before or at
substantially the same time as the second phases are brought into contact with the flow of
the first phase.
In one embodiment, the flow of the second phase is brought into contact with the flow of
the first phase substantially perpendicular to the first phase. In this embodiment, the
channel structure may be a T-junction geometry. The path of the channel may follow the
path of the flow of the first phase, in which case the second flow will be substantially
perpendicular to the resulting combined flow in the channel. Alternatively, the path of the
channel may follow the path of the flow of the second phase, in which case the first phase
flow will be substantially perpendicular to the resulting combined flow in the channel.
Methods utilising a T-junction geometry provide discrete regions, typically droplets, of the
aqueous phase in the oil phase as a result of induced shear forces within the two phase
system.
In one embodiment, an additional flow of the first phase is provided. The first phase flows
are brought into contact with each side of the second phase flow in a channel, and the
flow of phases is then passed through a region of the channel of reduced cross-section
(an orifice) thereby to generate a discrete region, preferably a droplet, of the second
phase in the channel. Such methods, which have an inner second phase flow and two
outer first phase flows, are referred to as flow-focussing configurations.
Methods using flow-focussing techniques provide discrete regions, typically droplets, of
the second phase in the first phase as a result of the outer first phase applying pressure
and viscous stresses to the inner second phase, thereby generating a narrow flow of that
phase. This narrowed flow then separates into discrete regions, typically droplets, at the
orifice or soon after the combined flow has passed through the orifice.
In one embodiment, the discrete region is a droplet.
In one embodiment, the discrete region is a slug.
After the discrete region is formed in the channel, the discrete region may be passed
along the channel to a collection area. The residence time of the discrete region in the
channel is not particularly limited. In one embodiment, the residency time is sufficient to
allow the shell to form.
As the discrete region is passed along the channel it may be subjected to a mixing stage
whereby the components of the discrete region are more evenly distributed around that
discrete region. In one embodiment, the channel comprises a winding region. The
winding region may take the form of a substantially sigmoid path through which the
discrete region is passed.
In one embodiment, the second phase further comprises a component for encapsulation,
and the step i) provides a capsule encapsulating the component.
The first phase and the second phase may be contacted at a simple T-junction. The
second phase may be formed from the combination of separate flows of cucurbiturii and
(1), (2) or (3) as appropriate. Where there are more than two components, these
components may be brought into contact simultaneously or sequentially.
These flows may be contacted prior to contact with the first phase. Alternatively they may
be brought into contact simultaneously on contact with the first phase.
Discrete regions of second phase are generated in the channel as the immiscible first
phase shears off the second phase. The frequency of shearing is dependent on the flow
rate ratio of the two phases.
In one embodiment, the flow rate is selected so as to provide a set number of droplets per
unit time (droplets per second).
The droplets may be prepared at a rate of at most 0,000, at most, 5,000, at most ,000
or at most 500 Hz.

Claims
. A capsule having a shell which is obtainable from the complexation of a
composition comprising a host and one or more building blocks having suitable host guest
functionality thereby to form a supramolecular cross-linked network.
2. The capsule of claim , wherein the host is selected from cucurbituril, cyclodextrin,
calix[n]arene, and crown ether, and the one or more building blocks have suitable host
guest functionality for the cucurbituril, cyclodextrin, calix[n]arene or crown ether host.
3. The capsule of claim 2, wherein the host is cucurbituril and the one or more
building blocks have suitable cucurbituril guest functionality.
4. The capsule of claim 3, wherein the shell is obtainable from the complexation of
(a) a composition comprising cucurbituril and ( 1 ) or (2); or (b) a composition comprising a
plurality of covaient!y linked cucurbituril and ( 1 ) , (2) or (3), wherein:
(1) comprises a first building block covalentiy linked to a plurality of first cucurbituril
guest molecules and a second building block covalentiy linked to a plurality of second
cucurbituril guest molecules, wherein a first guest molecule and a second guest molecule
together with cucurbituril are suitable for forming a ternary guest-host complex.
(2) comprises a first building block covalentiy linked to a plurality of first cucurbituril
guest molecules and a plurality of second cucurbituril guest molecules, wherein a first and
a second guest molecule together with cucurbituril are suitable for forming a ternary
guest-host complex and optionally the composition further comprises a second building
block covalentiy linked to one or more third cucurbituril guest molecules, one or more
fourth cucurbituril guest molecules or both, wherein a third and a fourth molecule together
with cucurbituril are suitable for forming a ternary guest-host complex, and/or the first and
fourth molecules together with cucurbituril are suitable for forming a ternary guest-host
complex, and/or the second and third molecules together with cucurbituril are suitable for
forming a ternary guest-host complex; and
(3) comprises a first building block covalentiy linked to a plurality of first cucurbituril
guest molecules, wherein the first guest molecule together with the cucurbituril are
suitable for forming a binary guest-host complex. Optionally the composition further
comprises a second building block covalentiy linked to one or more second cucurbituril
guest molecules, wherein the second guest molecule together with the cucurbituril are
suitable for forming a binary guest-host complex.
5. The capsule of claim 4 wherein the shell is obtainable from the complexation of a
composition comprising cucurbiturii and ( 1) or (2).
6. The capsule of claim 5, wherein the shell is obtainable from the complexation of a
composition comprising cucurbiturii and ( 1 ) .
7. The capsule of any one of claims 3 to 6, wherein the cucurbiturii is selected from
CB[8] and variants and derivatives thereof.
8. The capsule of claim 7 wherein the cucurbiturii is CB[8].
9. The capsule of claim 7 or claim 8, wherein the cucurbiturii forms a ternary complex
with a first guest molecule and a second guest molecule, and the first and second guest
molecules are selected from the following pairs:
viologen and naphthol;
viologen and dihydroxybenzene;
viologen and tetrathiafu!va!ene;
viologen and indole;
methyiviologen and naphthol;
methyiviologen and dihydroxybenzene;
methyiviologen and tetrathiafulvalene;
methyiviologen and indole;
/,/V-dimethy!dipyridy!iumylethyiene and naphthol;
/V,/V~dimethyidipyridyiiumylethyiene and dihydroxybenzene;
/S/,/V-dimethy!dipyridy!iumylethyiene and tetrathiafulvalene;
A/,/V-dimethyidipyridyiiumylethyiene and indole;
2,7-dimethyldiazapyrenium and naphthol;
2,7-dimethyldiazapyrenium and dihydroxybenzene;
2,7-dimethyldiazapyrenium and tetrathiafulvalene; and
2,7-dimethyldiazapyrenium and indole.
10. The capsule of any one of claims 1 to 9, wherein the first building block is a
polymeric molecule.
11. The capsule of claim 10, wherein the polymeric molecule is or comprises a
po!y(meth)acry!ate-, a polystyrene- and/or a poly(meth)acrylamide polymer.
12. The capsule of ciaim 10 or ciaim 1, wherein the polymeric molecule comprises a
detectable label.
3. The capsule of any one of claims 1 to wherein the second building block,
where present, is a particle.
4. The capsule of claim 13, wherein the particle is or comprises gold or silver or
mixtures thereof.
15. The capsule according to any one of claims 1 to 14, wherein the capsule size is in
range from 0 to 100 mhi in diameter.
6. The capsule according to any one of claims 1 to 15, wherein the capsule diameter
has a relative standard deviation (RSD) of at most 10%.
1 . The capsule according to any one of claims 1 to 16, wherein the shell pore size is
in range 1 to 20 n .
8. The capsule according to any one of claims 1 to 7, wherein the capsule
encapsulates a component.
19. The capsule according to claim 18, wherein the component is a biological
molecule.
20. A method for the preparation of a capsule having a shell, the method comprising
the step of:
(i) contacting a flow of a first phase and a flow of a second phase in a channel,
thereby to generate in the channel a dispersion of discrete regions, preferably droplets, of
the second phase in the first phase, wherein the second phase comprises cucurbituril and
one or more building blocks having suitable cucurbiturilguest functionality suitable to form
a supramolecular cross-linked network, thereby to form a capsule shell within the discrete
region, wherein the first and second phases are immiscible.
2 1. The method of claim 20, wherein the second phase either (a) a cucurbituril and ( )
or (2); or (b) a plurality of covalently linked cucurbituril and (1), (2) or (3), wherein:
(1) comprises a first building block covalently linked to a plurality of first cucurbituril
guest molecules and a second building block covalently linked to a plurality of second
cucurbituril guest molecules, wherein a first guest molecule and a second guest molecule
together with cucurbituril are suitable for forming a ternary guest-host complex.
(2) comprises a first building block covalently linked to a plurality of first cucurbituril
guest molecules and a plurality of second cucurbituril guest molecules, wherein a first and
a second guest molecule together with cucurbituril are suitable for forming a ternary
guest-host complex and optionally the composition further comprises a second building
block covaientiy linked to one or more third cucurbituril guest molecules, one or more
fourth cucurbituril guest molecules or both, wherein a third and a fourth molecule together
with cucurbituril are suitable for forming a ternary guest-host complex, and/or the first and
fourth molecules together with cucurbituril are suitable for forming a ternary guest-host
complex, and/or the second and third molecules together with cucurbituril are suitable for
forming a ternary guest-host complex; and
(3) comprises a first building block covaientiy linked to a plurality of first cucurbituril
guest molecules, wherein the first guest molecule together with the cucurbituril are
suitable for forming a binary guest-host complex. Optionally the composition further
comprises a second building block covaientiy linked to one or more second cucurbituril
guest molecules, wherein the second guest molecule together with the cucurbituril are
suitable for forming a binary guest-host complex.
22. The method of claim 2 1 wherein the second phase comprises cucurbituril and (1)
or (2).
23. The method of claim 22, wherein the second phase comprises cucurbituril and ( 1) .
24. The method of any one of claims 20 to 23, wherein the cucurbituril is selected from
CB[8] and variants and derivatives thereof.
25. The method of claim 24, wherein the cucurbituril is CB[8j.
26. The method of claims 24 or 25, wherein the cucurbituril forms a ternary complex
with a first guest molecule and a second guest molecule, and the first and second guest
molecules are selected from the following pairs:
vio!ogen and naphtho!;
vio!ogen and dihydroxybenzene;
viologen and tetrathiafu!va!ene;
viologen and indole;
methyivioiogen and naphthoi;
methylvioiogen and dihydroxybenzene;
methyivioiogen and tetrathiafulvaiene;
methyivioiogen and indole;
/V,/V-dimethyidipyridyiiumylethylene and naphthoi;
/,/V-dimethyidipyridyiiumylethyiene and dihydroxybenzene;
/V,/V-dimethy!dipyridy!iumylethyiene and tetrathiafulvaiene;
W,A/'-dimethyidipyridyiiumylethy!ene and indole;
2,7-dimethyldiazapyrenium and naphthoi;
2,7-dimethyldiazapyrenium and dihydroxybenzene;
2,7-dimethyldiazapyrenium and tetrathiafuivalene; and
2,7-dimethyldiazapyrenium and indole.
27 The method of any one of claims 20 to 26, wherein the first building block is a
polymeric molecule.
28. The method of claim 7, wherein the polymeric molecule is or comprises a
poiy(meth)acry!ate-, a polystyrene- and/or a poly(meth)acrylamide polymer.
29. The method of claim 27 or claim 28, wherein the polymeric molecule comprises a
detectable label.
30. The method of any one of claims 20 to 29, wherein the second building block,
where present, is a particle.
3 . The method of claim 30, wherein the particle is or comprises gold or silver or
mixtures thereof.
32. The method of any one of claims 20 to 3 , wherein the second phase is an
aqueous phase and the first phase is a water immiscible phase.
33. The method of any one of claims 20 to 32, wherein the second phase further
comprises a component for encapsulation, and the step (i) provides a capsule having a
shell encapsulating the component.
34. The method of any one of claims 20 to 33, wherein the method further comprises
the step of (ii) collecting the outflow from the channel, thereby to obtain a droplet, which
contains a capsule.
35. The method of claim 34, further comprising the step of drying the capsule obtained
in step (ii).
36. A method of delivering a component to a location, the method comprising the
steps of:
(i) providing a capsule having a shell encapsulating a component, as defined in
claim 18 or claim 19;
(ii) delivering the capsule to a target location;
(iii) releasing the component from the shell.